Synthesis of archaeal bipolar lipid analogues: A way to versatile drug/gene delivery systems

Article abstract of DOI:10.1021/jo071181r

(Chemical Equation Presented) A synthetic route for the preparation of symmetrical and unsymmetrical archaeal tetraether-like analogues has been described. The syntheses are based upon the elaboration of hemimacrocyclic tetraether lipid cores from versati

Full text of DOI:10.1021/jo071181r

Synthesis of Archaeal Bipolar Lipid Analogues: A Way to Versatile
Drug/Gene Delivery Systems
Mickae¨lle Brard, Ce´line Laine´, Gildas Re´thore´, Isabelle Laurent, Ce´cile Neveu,
Lo¨ıc Lemie`gre, and Thierry Benvegnu*
UMR CNRS 6226 “Sciences Chimiques de Rennes”, Equipe “Chimie Organique et Supramole´culaire”,
Ecole Nationale Supe´rieure de Chimie de Rennes, AV. Ge´ne´ral Leclerc, 35700 Rennes, France
thierry.benVegnu@ensc-rennes.fr
ReceiVed June 4, 2007
A synthetic route for the preparation of symmetrical and unsymmetrical archaeal tetraether-like analogues
has been described. The syntheses are based upon the elaboration of hemimacrocyclic tetraether lipid
cores from versatile building blocks followed by simultaneous or sequential introduction of polar head
groups. Functionalizations of the tetraether lipids with neutral lactose or phosphatidylcholine polar heads
and cationic glycine betaine moieties were envisaged both to increase membrane stability and to exhibit
interactions with charged nucleic acids. Additionally, mannose and lactose triantennary clusters designed
as multivalent ligands for selective interaction with lectin-type receptors were also efficiently synthesized
for active cell/tissue targeting.
Introduction
efficient in delivering the drug in a target-specific manner, stable
in vitro as well as in vivo, nontoxic, non-immunogenic, and it
should reduce the need for multiple administrations. Liposome
delivery systems answer most of these conditions, and numerous
studies have thus demonstrated the potentiality of liposomes as
carriersofdrugs,vaccines,genes,imagingagents,orantioxidants.1-6
However, a major drawback of liposome formulations is their
lack of stability all along the extracellular route to the target
sites, reducing the circulation time. Especially if an oral
administration is considered, their poor stability at low pH and
their tendency to be attacked by bile salts, gastrointestinal tract
enzymes, or lipoproteins have considerably limited the interest
Phospholipid liposomes or synthetic lipid-containing vesicles
are self-assembled colloidal particles resulting from the orga-
nization of amphiphilic molecules in aqueous media. The
hydrophilic head groups of the lipids forming liposomes are
oriented toward the aqueous environments present inside and
outside the liposomes, whereas the hydrophobic regions of the
lipids are sandwiched between the polar head groups and away
from the aqueous environments. Liposomes termed unilamellar
vesicles (ULV) are composed of a single bilayer, and multila-
mellar vesicles (MLV, onion-like structure) contain multiple
bilayers with an aqueous space separating the bilayers from one
to the other. As liposome structures induce the presence of both
hydrophobic and hydrophilic domains, they are able to entrap
both hydrophilic and hydrophobic molecules, which is particu-
larly attractive for encapsulation and drug delivery applications.
An ideal vehicle or vector in drug delivery should be highly
(1) Xiang, T.-X.; Anderson, B. D. AdV. Drug DeliVery ReV. 2006, 58,
1357-1378.
(2) Dass, C. R.; Choong, P. F. M. J. Controlled Release 2006, 113, 155-
163.
(3) Mozafari, M. R.; Flanagan, J.; Matia-Merino, L.; Awati, A.; Omri,
A.; Suntres, Z. E.; Singh, H. J. Sci. Food Agric. 2006, 86, 2038-2045.
(4) Lian, T.; Ho, R. J. Y. J. Pharm. Sci. 2001, 90, 667-680.
(5) Chonn, A.; Cullis, P. R. AdV. Drug DeliVery ReV. 1998, 30, 73-83.
(6) Yao, T.; Huang, L.; Zern, M. A. AdV. Drug DeliVery ReV. 1995, 17,
239-246.
* To whom correspondence should be addressed. Tel: (33) (0)2 23 23 80
60. Fax: (33) (0)2 23 23 80 46.
10.1021/jo071181r CCC: $37.00 © 2007 American Chemical Society
Published on Web 10/03/2007
J. Org. Chem. 2007, 72, 8267-8279
8267

Brard et al.
FIGURE 1. Typical basic structures of macrocyclic and hemimacrocyclic natural archaeal lipids.
of liposome systems in the delivery of medicines. Attempts to
improve their stability either by incorporation of high amounts
of cholesterol in the bilayer (improve mechanical properties,
such as an increased stretching elastic modulus, resulting in
stronger membranes and reduced permeability) or by coating
the liposome surface with hydrophilic polyethyleneglycol
polymers (sterically stabilized liposomes, called stealth lipo-
somes, to reduce interactions with blood proteins) have increased
the resident time and therefore have permitted better efficacy
and passive targeting. Nevertheless, these improvements have
not solved all the problems encountered with liposome technol-
ogy, notably insufficient in vivo stability and lack of active
targeting.
Archaeosomes are liposomes which are made from natural
lipids found in archaea or from synthetically derived lipids that
have the unique structure characteristics of archaeal membrane
lipids.7,8 The lipid components of archaea are strikingly different
than those found in other forms of life, and they are considered
as specific and useful markers of this evolutive line. Apart from
the archaea domain, all forms of life usually have lipids that
contain ester linking groups. However, for archaea, the lipids
contain ether linkages and saturated isopranyl units (Figure 1).
The ether linkages are chemically and enzymatically more stable
than esters located in comparable positions within the structures
of the lipids, and the branching methyl groups help to keep the
membrane in a fluid state. In addition to possessing saturated
branched residues and ether moieties, lipids of the archaea
domain can also possess aliphatic phytanyl chains ether-linked
to two sn-2,3-glycerol units that span the membrane from the
inner to the outer side. When two chains of a particular head
group span the membrane to another head group on the opposing
surface, a macrocyclic unit is formed (Figure 1, type A), and
when only one chain spans the membrane, a hemimacrocycle
is formed (Figure 1, type B). The monolayer-type membrane
organizations obtained from these tetraether derivatives inhibit
lipid lateral diffusion phenomena and reduce flip-flop dynamics
in comparison with standard bilayer systems.9 In some cases,
cyclopentyl units are incorporated into the spanning chains, and
their number (up to 8) usually increases with the roughness of
the life conditions.10,11
Thus, membranes of the archaea domain are relatively more
stable than those of other forms of life, thereby enabling
organisms containing membranes of this type to survive under
extreme conditions of pH, temperature, pressure, salt concentra-
tion, and anaerobia. Within this context, natural archaeal lipids
have been used as innovative materials in liposome formulations,
exploiting their ability to enhance lipid membrane stabilities.12,13
The poor availability of pure natural archaeal lipids forces
scientists to design and synthesize realistic analogues. Indeed,
approaches to synthetic hemimacrocyclic and macrocyclic
tetraethers related to archaeal membranes have been reported
during the past 20 years.14-22 These synthetic bipolar compounds
generally contain branched phytanyl chains and/or linear oli-
gomethylene bridging chains; in some cases, aromatic or
heterocyclic rings are also incorporated into the hydrophobic
core. However, among the number of analogues already
prepared so far, neither contains the specific cyclopentane rings
and only few synthetic compounds reach the same length as in
natural archaeal lipids. Additionally, most of the synthetic efforts
were directed toward the preparation of tetraethers bearing
phosphate moieties as polar head groups. No reports had
appeared on the synthesis of neutral glycosylated or cationic
tetraethers until our preliminary reports on the development of
lactose- or glycine betaine-containing tetraethers for the devel-
opment of drug or gene delivery systems.23,24 Our strategy aims
at fine-tuning the properties of conventional liposome mem-
(11) De Rosa, M.; Esposito, E.; Gambacorta, A.; Nicolaus, B.; Bu’Lock,
J. D. Phytochemistry 1980, 19, 827-831.
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357.
(13) Patel, G. B.; Chen, W. Curr. Drug DeliVery 2005, 2, 407-421.
(14) O’Neil, E. J.; DiVittorio, K. M.; Smith, B. D. Org. Lett. 2007, 9,
199-202.
(15) Benvegnu, T.; Brard, M.; Plusquellec, D. Curr. Opin. Colloid
Interface Sci. 2004, 8, 469-479.
(16) Brard, M.; Richter, W.; Benvegnu, T.; Plusquellec, D. J. Am. Chem.
Soc. 2004, 126, 10003-10012.
(17) Miyawaki, K.; Harada, A.; Takagi, T.; Shibakami, M. Synlett 2003,
349-352.
(18) Miyawaki, K.; Goto, R.; Takagi, T.; Shibakami, M. Synlett 2003,
1467-1470.
(19) Lecollinet, G.; Gulik, A.; Mackenzie, G.; Goodby, J. W.; Benvegnu,
T.; Plusquellec, D. Chem.sEur. J. 2002, 8, 585-593.
(20) Raguse, B.; Culshaw, P. N.; Prashar, J. K.; Raval, K. Tetrahedron
Lett. 2000, 41, 2971-2974.
(7) De Rosa, M.; Gambacorta, A.; Gliozzi, A. Microbiol. ReV. 1986,
70-80.
(21) Cuccia, L. A.; Morin, F.; Beck, A.; He´bert, N.; Just, G.; Lennox,
R. B. Chem.sEur. J. 2000, 6, 4379-4384.
(22) Heathcock, C. H.; Finkelstein, B. L.; Jarvi, E. T.; Radel, P. A.;
Hadley, C. R. J. Org. Chem. 1988, 53, 1922-1942.
(23) Benvegnu, T.; Re´thore´, G.; Brard, M.; Richter, W.; Plusquellec, D.
Chem. Commun. 2005, 5536-5538.
(8) Sprott, G. D. J. Bioenerg. Biomembr. 1992, 24, 555-566.
(9) Shinoda, W.; Shinoda, K.; Baba, T.; Mikami, M. Biophys. J. 2005,
89, 3195-3202.
(10) Tomoaia-Cotisel, M.; Chifu, E.; Zsako, J.; Mocanu, A.; Quinn, P.
J.; Kates, M. Chem. Phys. Lipids 1992, 63, 131-138.
8268 J. Org. Chem., Vol. 72, No. 22, 2007

Synthesis of Archaeal Bipolar Lipid Analogues
results from our aim to simplify the synthetic pathway initially
developed for tetraethers 1a, 2-4, 5a, 6, and 7.
Results and Discussion
Preparation of Tetraether Diols. Intrigued by the presence
of cyclopentane rings in natural archaeal lipids and their
influence on the membrane structure, we have investigated and
designed synthetic pathways that permit the introduction of five-
membered rings into the hydrophobic core (Figure 4). Actually,
the role of the cyclopentane residues in membrane properties
was modeled by Chong et al., who clearly showed significant
changes in the packing arrangement of structures containing such
rings or not (0 or 4 rings).25 These preliminary models denote
a thinner and more packed membrane for the tetraether
containing four rings than for the tetraether without any ring.
In the present study, we focused on a cis-configured 1,3-
disubstituted cyclopentyl unit; however, it is important to note
that Montenegro et al. determined recently, by synthesis and
NMR comparisons, the cyclopentane configuration in a natural
archaeal lipid to be trans.26 Our synthetic strategy is based on
a final coupling between a phytanylated glycerol 8 and a
symmetrical cyclopentane-containing bridging chain 9a,b (Fig-
ure 4). Building block 8 was synthesized from phytol 10 and
(R)-solketal 11, and spacers 9a,b were prepared from diol 12,
which results from an ozonolysis/reduction sequence on nor-
bornene 13.
FIGURE 2. Stabilization of conventional liposome via the introduction
of tetraether lipids.
branes (rigidity, fluidity) via the controlled modulation of its
lipid composition (bilayer/monolayer-forming lipids) as natural
archaea do. This work takes advantage of the outstanding
properties of archaeal lipids to improve both the stability of
the delivery system and the anchoring of cell-surface ligands
into the lipid membrane (Figure 2).
In this paper, we report full details of the preparation of
symmetrical and unsymmetrical neutral or cationic tetraether-
type compounds 1-5 (Figure 3). Furthermore, with the aim to
develop site-specific drug targeting, novel unsymmetrical
synthetic tetraethers 6 and 7 containing a triantennary sugar
(lactose or mannose) moiety were designed and synthesized in
order to bind to lectin-type receptors (over)expressed on the
cell surfaces.
FIGURE 4. Retrosynthesis scheme tetraether analogues 1a,b.
FIGURE 3. Synthetic analogues of archaeal membrane bipolar lipids.
The results on the preparation of the glyceryl alcohol 8 have
been previously reported;20,27,28 however, due to low regiose-
lectivity and/or insufficient efficiency of the syntheses, we
improved its preparation from (R)-solketal 11 and phytol 10.
In brief, commercially available (R)-solketal 11 was benzylated
(BnBr, NaH) at the primary position, and the acetonide moiety
was cleaved under acidic conditions (IR-120, MeOH) to afford
the diol 14 (85%, two steps) (Scheme 1). The latter was then
monoprotected with trityl chloride (68% yield), and 15 reacted
with phytanyl bromide29 to furnish the differentially protected
The synthetic hemimacrocyclic bipolar lipids are actually
characterized by the following: (1) the presence of a 31 or 33
atom-long bridging chain containing a cyclopentane ring (sup-
posed to increase the lipid dispersion into water)16 linked at
both ends to two (S)-glycerol moieties via ether bonds, (2) two
phytanyl chains having a combined length equivalent to that of
the bridging chain, and (3) neutral hydroxyl groups (lipids 1a,b),
lactosyl moieties (lipid 2), phosphatidylcholine units (lipids 3
and 4), positive quaternary ammonium groups derived from
glycine betaine to provide electrostatic interactions with nega-
tively charged DNA (lipids 5a,b), or a hydroxyl unit and a
trimannosylated or trilactosylated ligand introduced at opposite
ends of the lipophilic backbone (6 and 7). The presence of
oxygen atoms within the spanning chains of lipids 1b and 5b
(25) Gabriel, J. L.; Lee Gau Chong, P. Chem. Phys. Lipids 2000, 105,
193-200.
(26) Montenegro, E.; Gabler, B.; Paradies, G.; Seemann, M.; Helmchen,
G. Angew. Chem., Int. Ed. 2003, 42, 2419-2421.
(27) Eguchi, T.; Arakawa, K.; Kakinuma, K.; Rapp, G.; Ghosh, S.;
Nakatani, Y.; Ourisson, G. Chem.sEur. J. 2000, 6, 3351-3358.
(28) Aoki, T.; Poulter, C. D. J. Org. Chem. 1985, 50, 5634-5636.
(29) Yamauchi, K.; Kinoshita, M. Prog. Polym. Sci. 1993, 18, 763-
804.
(24) Re´thore´, G.; Montier, T.; Gall, T. L.; Dele´pine, P.; Cammas-Marion,
S.; Lemie`gre, L.; Lehn, P.; Benvegnu, T. Chem. Commun. 2007, 2054-
2056.
J. Org. Chem, Vol. 72, No. 22, 2007 8269

Brard et al.
SCHEME 2. Synthesis of Archaeal Lipid Analogues 1a,b
SCHEME 1. Preparation of Glyceryl Derivative 8
monophytanyl glycerol derivative. Final cleavage of the trityl
group with iron chloride hexahydrate provided the targeted
building block 8 in good yield (70%).30 At this stage, we have
been able to scale-up the synthesis of compound 8 to a 40 g
scale with an overall yield of 35% (five steps).
The introduction of two oxygen atoms into the middle of the
lipidic skeleton (1b) is an important structural difference, and
the corresponding analogues dissent from natural archaeal lipids.
However, our choice results from an easier synthetic route
developed for the preparation of these tetraether-type analogues.
Indeed, the bridging chain (9b) of compound 1b was synthesized
via a more efficient and a shorter reaction sequence than for its
counterpart lipid 1a, starting from the same diol 1216 (Scheme
2). Treatment of diol 12 with triphenylphosphine and iodine
led to the formation of the corresponding 1,3-bis(iodomethyl)-
cyclopentane, which underwent a nucleophilic displacement with
triphenylphosphine in acetonitrile, providing the diphosphonium
salt 16 in 50% yield (two steps).31 Addition of 2 equiv of nBuLi
conduced to the corresponding diylide that performed a double
Wittig olefination with 12-benzyloxydodecancarboxaldehyde.
A subsequent pallado-catalyzed hydrogenation of the resulting
dialkene furnished in a one-pot reaction the saturated and
deprotected diol 9a in 30% overall yield (four steps from diol
12). Coupling of 2 equiv of 12-benzyloxydodecanetriflate with
diol 12 in the presence of proton sponge (PS) followed by a
pallado-catalyzed hydrogenolysis of the benzyloxy groups
provided the hydrophobic chain 9b. The quality of the triflate
anhydride was found to have a dramatic influence on the
reaction yield. The best yield achieved for this two-step sequence
was 80%, with an average of 65% for repeated runs. In addition,
replacement of proton sponge by hydride bases such as NaH
or KH was found to be ineffective and led mainly to elimination
byproducts. The use of mesylated compounds also failed
whatever the base used. Treatment of alcohol 8 with KH at 0 °C
in dry THF followed by coupling with the ditriflates of diols
9a,b afforded dibenzylated tetraethers 17a,b in 64 and 45%
yield, respectively. A final hydrogenolysis of the benzyloxy
groups was conduced in the presence of palladium on carbon
and concretized the preparation of our archaeal lipid analogues
1a and 1b in overall yields of 15 (seven steps) and 26% (five
steps), respectively. We were able at this stage to scale-up the
preparation of both bipolar lipids 1a and 1b to multigram
quantities. However, concerning the preparation of 1a, optimized
yields were obtained when performing multiple batches of the
double Wittig olefination instead of a unique large scale batch.
Introduction of Polar Heads. Having the diols 1a,b in hand,
our next goal was to introduce various polar head groups at
one end or at both ends of these hemimacrocyclic structures.
Indeed, to fine-tune the properties of our archaeal-like lipids,
we envisaged further functionalizations of diols 1a,b with
neutral, zwitterionic, or cationic moieties. A series of mono-
and difunctionalized tetraethers were thus prepared from diol
1a (Scheme 3). Sugar polar heads are common in archaeal
membrane lipids, and they exert tremendous influence on the
membrane stability through highly cooperative polar interactions
between polyol heads. Thus, we investigated the introduction
of disaccharidic lactosyl groups in order to bring a sufficient
hydrophilic character to our synthetic glycolipids. A double
glycosylation reaction was carried out under standard thiogly-
coside chemistry32 (NIS, Et3SiOTfcat) and proceeded at 0 °C to
provide exclusively â-linked bislactosyl lipids. The â-selectivity
resulted from neighboring group participation of the C-2 acetyl
donor functionality. A final deprotection of the hydroxyl groups
by a Zemple`n procedure (MeONa, MeOH) provided the fully
deprotected dilactosyl tetraether 2 in 32% yield (two steps). The
introduction of two phosphatidylcholine groups was performed
via the double condensation of diol 1a on bromomethyl
phosphorodichloridate at room temperature, providing the
corresponding diphosphonate, which reacts subsequently with
(30) Ding, X.; Wang, W.; Kong, F. Carbohydr. Res. 1997, 303, 445-
448.
(31) Pohnert, G.; Boland, W. Eur. J. Org. Chem. 2000, 1821-1826.
(32) Fraser-Reid, B.; Udodong, U. E.; Wu, Z.; Ottosson, H.; Merritt, J.
R.; Rao, C. S.; Roberts, C.; Madsen, R. Synlett 1992, 927-942.
8270 J. Org. Chem., Vol. 72, No. 22, 2007

Synthesis of Archaeal Bipolar Lipid Analogues
SCHEME 3. Introduction of Phosphatidylcholine, Lactose, Benzyl, and Acetyl Groups onto Diol 1a
trimethylamine in 4 days to give the diphosphatidylcholine
tetraether 3 in 25% yield (two steps). Interestingly, the reaction
of an excess of 2-chloro-2-oxo-1,2,3-dioxaphospholane33-35
provided a selective monofunctionalization of diol 1a. The
following condensation of trimethylamine at 65 °C afforded the
corresponding monophosphatidylcholine tetraether 4 in a rela-
tively good yield (46%, two steps). Except for the latter example,
the efficient monofunctionalization of 1a required a selective
protection of one of the two hydroxyl groups. We therefore
investigated two types of monoprotection reaction (benzylation
or acetylation). The benzyl bromide/NaH system provided clean
monobenzylated tetraether 18 (51% yield), whereas monoacety-
lation of diol 1a with acetic anhydride in the presence of
triethylamine afforded monoacetylated tetraether 19 in 49% yield
(recyclable diacetylated tetraether and remaining diol 1a were
isolated in 33 and 17% yield, respectively).
These bipolar lipids are supposed to exhibit properties in
terms of membrane stability and permeability similar to those
of lipids from natural thermoacidophile organisms.29,34,36-38 In
a previous study, these synthetic analogues were found to
provide stable liposomes over wide temperature, in acidic
conditions (pH 2) or/and in the presence of fetal calf serum
(FCS) or sodium cholate (reflecting a bile salt-containing
media).23 In particular, incorporation of 30% of dilactosyl
tetraether 2 into egg PC liposome formulations remarkably
lowered the leakage of encapsulated 5(6)-carboxyfluorescein
(CF) in 0.4% sodium cholate and serum (FCS) media. Ad-
ditionally, archaeosomes made from exclusively diPC tetraether
3 have a pH stability that compares favorably to the stability of
liposomes composed of natural Thermoplasma acidophilium
tetraether lipids, which lost 20-30% of the marker after 10 min
at pH 2.
Another application of archaeosomes as innovative gene
delivery systems was additionally envisaged via the use of new
synthetic cationic tetraether lipids. The prerequisite for gene
therapy is carrier systems that envelop the therapeutic nucleic
acid and facilitate incorporation by the target cell. This special
problem of drug delivery needs to take into account the special
properties of negatively charged nucleic acids. Typically,
cationic lipids can be used to complex nucleic acids, thereby
forming so-called lipoplexes; these complexes are then able to
deliver genes into cells.39-41 Numerous combinations of DNA
with suspensions of vesicles composed of cationic lipids and
neutral “helper” lipids, such as DOPE (dioleoylphosphatidyle-
thanolamine) or cholesterol, were developed for transfection,
but their use in clinical gene therapy trials was relatively
unsatisfactory. Within this context, dicationic tetraethers 5a,b
were thus prepared from diols 1a,b by introducing glycine
betaine groups. Glycine betaine is a quaternary ammonium salt
representing an important byproduct of the sugar industry (27%
w/w of the molasses of sugar beet), and for this reason, it can
be regarded as an interesting renewable raw material (Scheme
4).42 A three-step sequence involving (1) a mesylation of the
diols 1 by mesyl chloride, (2) a condensation of sodium azide
in DMF at reflux, and (3) a pallado-catalyzed hydrogenation in
EtOH/THF 1:1 provided diamines 20a,b in 76 and 77% overall
yields, respectively (three steps). N-Acylations of the tetraether
diamines 20 were efficiently performed with the N-acyl thia-
zolidine-2-thione derivative 2143 of glycine betaine, affording
dicationic tetraethers 5a,b in 80% yield.
The reduction of the diazides to diamines 20 needs more
comments. Indeed, when this step was done in a mixture of
chlorinated solvents and ethanol (CHCl3 or CH2Cl2/EtOH 1:1),
a side reaction was encountered consisting of a monochlorination
(33) Menger, F. M.; Chen, X. Y.; Brocchini, S.; Hopkins, H. P.; Hamilton,
D. J. Am. Chem. Soc. 1993, 115, 6600-6608.
(34) Yamauchi, K.; Sakamoto, Y.; Moriya, A.; Yamada, K.; Hosokawa,
T.; Higuchi, T.; Kinoshita, M. J. Am. Chem. Soc. 1990, 112, 3188-3191.
(35) Yamauchi, K.; Moriya, A.; Kinoshita, M. Biochim. Biophys. Acta
1989, 1003, 151-160.
(36) Komatsu, H.; Chong, P. L. G. Biochemistry 1998, 37, 107-115.
(37) Patel, G. B.; Agnew, B. J.; Deschatelets, L.; Fleming, L. P.; Sprott,
G. D. Int. J. Pharm. 2000, 194, 39-49.
(38) Gliozzi, A.; Relini, A.; Chong, P. L.-G. J. Membr. Sci. 2002, 206,
131-147.
(39) Gao, X.; Huang, L. Gene Ther. 1995, 2, 710-722.
(40) Miller, A. D. Angew. Chem., Int. Ed. 1998, 37, 1768-1785.
(41) Martin, B.; Sainlos, M.; Aissaoui, A.; Oudrhiri, N.; Hauchecorne,
M.; Vigneron, J. P.; Lehn, J.-M.; Lehn, P. Curr. Pharm. Des. 2005, 11,
375-394.
(42) Antoine, J. P.; Marcilloux, J.; Plusquellec, D.; Benvegnu, T.;
Goursaud, F.; Patent FR2869910 31p, 2005.
(43) Floch, V.; Legros, N.; Loisel, S.; Guillaume, C.; Guilbot, J.;
Benvegnu, T.; Ferrieres, V.; Plusquellec, D.; Ferec, C. Biochem. Biophys.
Res. Commun. 1998, 251, 360-365.
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Brard et al.
SCHEME 4. Preparation of Dicationic Archaeal Lipid
Derivatives 5a,b
FIGURE 5. Freeze-fracture electron micrographs of cationic liposomes
formed by (a) lipid 5a and (b) lipid 5a/DOPE (95:5 w/w).
SCHEME 6. Synthesis of Trimannosyl and Trilactosyl
Clusters
SCHEME 5. Chlorination/Reduction of Diazide 22a
of the diazide 22a simultaneously with the reduction of the
second azide. The subsequent acylation with 21 conduced to
the corresponding monocationic tetraether 23 (Scheme 5).
From a physicochemical point of view, we studied the
membrane organization of liposomes containing solely lipid 5a
or a lipid 5a/DOPE (95:5 w/w) mixture by freeze fracture
electron microscopy (FFEM), with the samples being etched
before shadowing in order to better visualize the fracture
propagation path. Interestingly, dispersion of bipolar lipid 5a
yielded at room temperature vesicles which were not fractured
in the usual way along the midplane of the membrane (Figure
5). As previously described for diPC tetraether 3,23 the absence
of a fracture midplane clearly indicates that the bipolar lipids
span the membrane, forming a monolayer as with natural
tetraethers (Figure 5a). The addition of 5% of DOPE yielded a
small number of vesicles with midplane-fractured membranes
(Figure 5b). Nevertheless, even in the presence of the bilayer-
forming DOPE, the predominant structures observed were small
cross-fractured vesicles due to the membrane-spanning tetraether
lipids.
Preliminary studies clearly revealed that combinations of
DNA with suspensions of vesicles composed of these cationic
tetraethers 5a,b and neutral helper lipids such as DOPE exhibited
efficient in vitro gene transfection properties. Cationic archaeo-
somes represent a new approach for modulating the lipidic
membrane rigidity of the complexes they form with DNA.24
Targeting Archaeal Lipids. Lactose and mannose glyco-
conjugates are known to have specific interactions with lectin-
type receptors and thus are good candidates for active targeting
delivery. Lactosylated polymeric micelles exhibited higher
transfection efficiency against cultured HepG2 cells possessing
the asialoglycoprotein receptor in comparison to nontargeted
micelles due to the contribution of a receptor-mediated endocy-
tosis mechanism.44 In the same way, mannosylated liposomes
were recognized by macrophages via the mannose receptor and
(44) Wang, S.-n.; Deng, Y.-h.; Xu, H.; Wu, H.-b.; Qiu, Y.-k.; Chen,
D.-w. Eur. J. Pharm. Biopharm. 2006, 62, 32-38.
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Synthesis of Archaeal Bipolar Lipid Analogues
SCHEME 7. Introduction of Trimannosyl and Trilactosyl Clusters on Tetraether Lipids
led to specific biodistribution profiles.45 Dilactosyl tetraether 2
may interact with cell surface; however, the promiscuity between
the lipid and the ligand would probably inhibit interactions with
the targeted cells. We thus designed elongated triglycosyl-type
clusters which are known to increase significantly cell
recognition.46-48 These clusters are based on a pentaerythritol
skeleton which is particularly adapted to that purpose. Monoben-
zylation of pentaerythritol (24) was achieved through a one-
step alkylation with 0.25 equiv of benzyl bromide in the
presence of tetrabutylammonium iodide and NaH (0.28 equiv).49
The triallylation of the resulting triol with allyl bromide gave
compound 2550 in good yield (81%) (Scheme 6). Hydroboration
reactions with 9-BBN followed by the in situ conversion to the
corresponding triol under oxidative/basic conditions provided
triol 26 (90% yield).51 Triglycosylation under standard glyco-
sylation conditions with readily available trichloroacetamidate
derivatives 27a,b52,53 permitted an easy access to trimannoside
R-anomers and trilactoside â-anomers 28a,b in 90 and 95%
yields, respectively. The reactions were promoted by TMSOTf
in CH2Cl2 at room temperature and needed the use of an excess
of the donors (20 equiv). Finally, hydrogenolysis of the
benzyloxy groups provided alcohols 29a,b in good yields (91
and 90%, respectively).
Trimannosyl and trilactosyl clusters 29a,b were then con-
verted into hemimethylthioacetals 30a,b in the presence of
dimethyl sulfide and benzoyl peroxide in acetonitrile (Scheme
7). The two partners 30a or 30b and monoacetylated tetraether
19 were thus ready to accomplish the final coupling. The
introduction of the sugar clusters on 19 was therefore achieved
in the presence of N-iodosuccinimide and catalytic triflic acid.
The final deacylation of all hydroxyl groups under Zemple`n
conditions (MeONa, MeOH) furnished the two targeting lipids
6 and 7 in quite fair yields (60%, three steps). The trimannosyl
tetraether 6 was easily and fully characterized by NMR (1H and
13C) and mass spectrometry. However, compound 7 was found
(45) Liang, H.-F.; Chen, C.-T.; Chen, S.-C.; Kulkarni, A. R.; Chiu, Y.-
L.; Chen, M.-C.; Sung, H.-W. Biomaterials 2006, 27, 2051-2059.
(46) Lee, Y. C.; Lee, R. T. Acc. Chem. Res. 1995, 28, 321-327.
(47) Frisch, B.; Carriere, M.; Largeau, C.; Mathey, F.; Masson, C.;
Schuber, F.; Scherman, D.; Escriou, V. Bioconjugate Chem. 2004, 15, 754-
764.
(48) Remy, J.; Kichler, A.; Mordvinov, V.; Schuber, F.; Behr, J. Proc.
Natl. Acad. Sci. U.S.A. 1995, 92, 1744-1748.
(49) Lehmann, J.; Weitzel, U. P. Carbohydr. Res. 1996, 294, 65-94.
(50) Daniellou, R. Ph.D. Thesis, Orsay, France, 2003, No. Ordre 7361.
(51) Hanessian, S.; Prabhanjan, H.; Qiu, D.; Nambiar, S. Can. J. Chem.
1996, 74, 1731-1737.
(52) Mei, X.; Heng, L.; Fu, M.; Li, Z.; Ning, J. Carbohydr. Res. 2005,
340, 2345-2351.
(53) Al-Mughaid, H.; Grindley, T. B. J. Org. Chem. 2006, 71, 1390-
1398.
J. Org. Chem, Vol. 72, No. 22, 2007 8273

Brard et al.
added, and the reaction mixture was stirred at room temperature
for 1.5 h. Five percent aqueous HCl was added, and the aqueous
phase was extracted with dichloromethane. The combined organic
phases were successively washed with saturated NaHCO3 and brine,
dried (MgSO4), and concentrated under reduced pressure. The
residue was purified by flash chromatography on silica gel (PE/
EtOAc 9:1) to yield corresponding 12-benzyloxydodecanetriflate
(6.05 g, 83%) as a brown oil. Due to its relative instability, the
compound was not characterized and was used rapidly in the next
step.
to be insoluble in most of common organic solvents but d5-
pyridine in which H NMR spectra could be easily recorded.
1
The structure of 7 was also confirmed by high-resolution mass
spectrometry. This drawback could make its formulation less
easy than for its mannosyl counterpart, but it would not prevent
the formation of archaeosomes including few percents of this
trilactosyl tetraether.
Conclusions
A mixture of diol 12 (400 mg, 14.3 mmol, 1.1 equiv) and proton
sponge (1.98 g, 9.2 mmol, 3 equiv) in dry dichloromethane (50
mL) was stirred at room temperature for 1 h. A solution of fresh
12-benzyloxydodecanetriflate in dry dichloromethane (10 mL) was
added, and the reaction mixture was stirred at reflux for 5 days.
The solvent was evaporated under reduced pressure, and the residue
was purified by flash chromatography on silica gel (PE/EtOAc 96:
4) to yield benzylated product (1.58 g, 79%) as a white solid: Rf
) 0.2 (PE/EtOAc 8:2); 1H NMR (CDCl3, 400 MHz) δ (ppm) 0.90-
0.98 (dt, J ) 9.1, 12.7 Hz, 1H), 1.23-1.38 (m, 34H), 1.50-1.58
(m, 8H), 1.68-1.78 (m, 2H), 1.91-1.99 (m, 1H), 2.13-2.24 (m,
2H), 3.28 (d, J ) 7.1 Hz, 4H), 3.39 (t, J ) 6.6 Hz, 4H), 3.62 (t, J
) 6.6 Hz, 4H), 4.50 (s, 4H), 7.25-7.37 (m, 10H); 13C NMR
(CDCl3, 100 MHz) δ (ppm) 26.59, 26.60, 29.3, 29.90, 29.91, 30.0,
30.1, 30.2, 34.3, 40.1, 70.9, 71.6, 73.2, 76.0, 127.9, 128.0, 128.8,
139.1; HRMS (ESI+) calcd for C45H74O4 (M + Na)+ 701.5485,
found 701.5490; (M + K)+ 717.5224, found 717.5227. Anal. Calcd
for C45H74O4: C, 79.59; H, 10.98. Found: C, 79.99; H, 10.90.
A mixture of previous dibenzylated diol (2.7 g, 4 mmol, 1 equiv)
in THF/MeOH (100 mL, 3:1) and palladium dihydroxide (700 mg,
5% w/w) was stirred at room temperature for 5 h under hydrogen
atmosphere. The reaction mixture was warmed (40-45 °C) and
filtered on Celite. The filtrate was concentrated under reduced
pressure, and the residue was recrystallized from cyclohexane to
yield 9b (1.7 g, 90%) as a white solid: Rf ) 0.2 (PE/EtOAc 8:2);
mp 63 °C; 1H NMR (CDCl3, 400 MHz) δ (ppm) 0.90-0.98 (dt, J
) 9.1, 12.7 Hz, 1H), 1.23-1.38 (m, 34H), 1.50-1.58 (m, 10H),
1.68-1.78 (m, 2H), 1.91-1.99 (m, 1H), 2.13-2.24 (m, J ) 6.6
Hz, 2H), 3.28 (d, J ) 7.1 Hz, 4H), 3.39 (t, J ) 6.6 Hz, 4H), 3.63
(t, J ) 6.6 Hz, 4H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 26.1,
26.6, 29.2, 29.8, 29.8, 29.96, 29.98, 30.1, 33.2, 34.3, 40.1, 63.5,
71.5, 76.0; HRMS (ESI+) calcd for C31H62O4 (M + Na)+ 521.4546,
found 521.4542. Anal. Calcd for C31H62O4: C, 74.64; H, 12.53.
Found: C, 74.61; H, 12.20.
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methylene]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-sn-diglycer-
ol 1a. Triflic anhydride (611 µL, 3.6 mmol, 3.8 equiv) was added
dropwise to a solution of 2,6-lutidine (423 µL, 7.6 mmol, 3.8 equiv)
in dry dichloromethane (20 mL) at 0 °C, and the resulting red
solution was stirred at 0 °C for 15 min. The diol 9a (500 mg, 0.96
mmol, 1 equiv) was added, and the reaction mixture was stirred at
room temperature and then at 30 °C until the complete dissolution
of 9a. Water was added, and the aqueous phase was extracted with
dichloromethane. The combined organic phases were successively
washed with 5% aqueous HCl, 5% aqueous NaHCO3 and brine,
dried (MgSO4), and concentrated under reduced pressure. The
residue was purified by flash chromatography on silica gel (PE/
EtOAc 9:1) to yield the corresponding ditriflate as a white solid:
Rf ) 0.8 (PE/EtOAc 9:1). Due to its relative instability the
compound was not further characterized and was rapidly used in
the next step.
In summary, we have synthesized a series of functionalized
hemimacrocyclic symmetrical or unsymmetrical tetraethers
where a cyclopentane ring is incorporated into the middle of
the bridging chain, which reminds one of the structure of
thermoacidophile archaeal lipids. The preparation of tetraether-
type diols 1a,b could be scaled up to multigram quantities that
permit one to envisage numerous functionalizations and potential
uses in drug/gene delivery. The introduction of various polar
head groups (lactose, mannose, phosphatidylcholine, glycine
betaine) provides a new generation of highly stable liposomes.
Tetraether 1a was also coupled with lactose and mannose
triantennary clusters for the development of targeting archaeo-
somes. The efficiency and flexibility of the synthetic routes
allow us to envisage the optimization of the ligand/receptor
interaction by modulating the length and the molecular structure
of the cluster between the lipidic core and the ligand.
Experimental Section
(cis-16,19-Methylidene)tetratriaconta-1,33-diol 9a. Wittig Ole-
fination. To a suspension of phosphonium salt 16 (300 mg, 0.34
mmol, 1 equiv) in dry THF (6 mL) was added a solution of
n-butyllithium (360 µL, 0.72 mmol, 2.1 equiv) in hexane (2 M) at
0 °C. The resulting orange mixture was stirred at 0 °C for 15 min.
A solution of 12-benzyloxydodecancarboxaldehyde (239 mg, 0.69
mmol, 2.0 equiv) in THF (6 mL) was added, and the reaction
mixture was stirred until discoloration. Water was added, and the
aqueous phase was extracted with (PE/EtOAc 6:4). The combined
organic phases were washed with water, dried (MgSO4), and
concentrated under reduced pressure. The residue was purified by
flash chromatography on silica gel (PE/EtOAc 99:1) to yield
unsaturated and dibenzylated product (150 mg, 63%) as a white
1
solid: Rf ) 0.6 (PE/EtOAc 8:2); H NMR (CDCl3, 400 MHz) δ
(ppm) (major isomer) 0.95-1.04 (m, 1H), 1.25-1.40 (m, 42H),
1.57-1.63 (m, 4H), 1.74-2.00 (m, 3H), 2.03 (m, 4H), 2.78 (m,
2H), 3.46 (t, J ) 6.6 Hz, 4H), 4.50 (s, 4H), 5.27-5.37 (m, 4H),
7.24-7.34 (m, 10H); 13C (CDCl3, 100 MHz) δ (ppm) (major
isomer) 26.2, 27.5, 29.3, 29.5, 29.6, 29.7, 29.8, 30.0, 32.8, 38.2,
42.2, 70.5, 72.8, 127.4, 126.6, 128.3, 128.7, 135.0, 138.7. Anal.
Calcd for C49H78O2: C, 84.18; H, 11.25. Found: C, 83.70; H, 10.84.
Hydrogenation. A mixture of the previous compound (1.93 g,
2.8 mmol, 1 equiv) and palladium on activated carbon (500 mg,
25% w/w) in EtOH (150 mL) was stirred overnight at room
temperature under hydrogen atmosphere. The reaction mixture was
warmed (40-45 °C) and filtered on Celite. The filtrate was
concentrated under reduced pressure to yield 9a (1.34 g, 93%) as
a white solid: Rf ) 0.4 (PE/EtOAc 6:4); mp 89-92 °C; 1H NMR
(CDCl3, 400 MHz) δ (ppm) 0.67 (m, 1H), 1.29 (m, 54H), 1.59 (m,
6H), 1.76 (m, 4H), 1.92 (m, 1H), 3.66 (t, J ) 6.6 Hz, 4H); 13C
(CDCl3, 100 MHz) δ (ppm) 25.9, 28.8, 29.6, 29.70, 29.72, 29.78,
29.81, 30.1, 31.9, 33.0, 36.9, 39.0, 40.4, 40.9, 63.2.
To a suspension of potassium hydride (438 mg, 3.8 mmol, 4
equiv) and alcohol 8 (1.32 g, 2.87 mmol, 3 equiv) in dry THF (12
mL) was added a solution of ditriflate (750 mg, 0.96 mmol, 1 equiv)
in dry THF (6 mL) at 0 °C. The reaction mixture was stirred at
0 °C for 10 min. Water was added, and the aqueous phase was
extracted with Et2O. The combined organic phases were dried
(MgSO4) and concentrated under reduced pressure. The residue was
purified by flash chromatography on silica gel (PE/EtOAc 10:0 then
cis-1,3-Bis((dodecan-1-ol)oxymethyl)cyclopentane 9b. Triflic
anhydride (5.67 mL, 34.2 mmol, 2 equiv) was added dropwise to
a solution of 2,6-lutidine (3.98 mL, 34.2 mmol, 2 equiv) in dry
dichloromethane (50 mL) at 0 °C. The resulting red solution was
stirred at 0 °C for 15 min. A solution of 12-benzyloxydodecanol
(5 g, 17.1 mmol, 1 equiv) in dry dichloromethane (20 mL) was
8274 J. Org. Chem., Vol. 72, No. 22, 2007

Synthesis of Archaeal Bipolar Lipid Analogues
95:5) to yield 17a (860 mg, 64%) as a yellow oil: Rf ) 0.8 (PE/
pressure. The residue was purified by flash chromatography on silica
gel (PE/EtOAc 9:1 then 7:3) to yield the peracetylated dilactosyl
tetraether (15 mg, 36%) as a gum: Rf ) 0.4 (PE/EtOAc 6:4); H
1
EtOAc 9:1); [R]20 +2.5 (c 0.81, CHCl3); H NMR (CDCl3, 400
D
1
MHz) δ (ppm) 0.59-0.64 (m, 1H), 0.83-1.07 (m, 30H), 1.07-
1.78 (m, 110H), 1.88 (m, 1H), 3.38-3.66 (m, 18H), 4.55 (s, 4H),
7.26-7.34 (m, 10H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 19.7,
19.77, 19.84, 22.7, 22.8, 24.5, 24.6, 24.9, 26.2, 28.1, 28.9, 29.6,
29.7, 29.8, 29.9, 30.1, 31.7, 32.9, 36.8, 37.16, 37.24, 37.4, 37.47,
37.54, 37.59, 38.9, 39.4, 40.2, 40.8, 69.0, 70.4, 70.8, 71.7, 73.4,
78.0, 127.6, 127.7, 128.4, 138.5. Anal. Calcd for C95H174O6: C,
80.79; H, 12.42. Found: C, 81.19; H, 12.50.
NMR (CDCl3, 400 MHz) δ (ppm) 0.51-0.59 (m, 1H), 0.71-0.81
(m, 30H), 1.00-1.65 (m, 110H), 1.81-1.85 (m, 1H), 1.90-2.09
(s, 42H), 3.31-3.52 (m, 18H), 3.72 (t, J ) 9.2 Hz, 2H), 3.79-
3.82 (m, 4H), 4.00-4.08 (m, 6H), 4.39-4.42 (m, 4H), 4.47 (d, J
) 7.9 Hz, 2H), 4.82-4.90 (m, 4H), 5.04 (dd, J ) 7.9, 10.4 Hz,
2H), 5.12 (t, J ) 9.4 Hz, 2H), 5.28 (d, J ) 2.3 Hz, 2H); 13C NMR
(CDCl3, 100 MHz) δ (ppm) 19.65, 19.70, 19.76, 19.83, 20.6, 20.7,
20.8, 20.9, 21.0, 22.7, 22.8, 24.5, 24.6, 24.9, 26.2, 28.1, 28.9, 29.6,
29.65, 29.73, 29.76, 29.82, 29.9, 30.0, 30.1, 31.7, 32.9, 36.8, 37.4,
37.46, 37.49, 37.54, 37.7, 38.8, 39.4, 40.2, 40.8, 60.9, 62.1, 66.7,
69.15, 69.23, 69.4, 70.5, 70.7, 71.1, 71.7, 71.9, 72.6, 73.0, 76.4,
77.9, 78.0, 100.9, 101.2, 169.2, 169.6, 169.9, 170.2, 170.3, 170.46,
170.47.
Deacetylation. A solution of previous peracetylated derivative
(15 mg, 0.006 mmol, 1 equiv) in MeOH/MeONa (2 mL, 0.05 M)
was stirred at room temperature for 2 h. A solution of acetic acid
in MeOH was added, and the reaction mixture was concentrated
under reduced pressure. The residue was purified on LH-20
Sephadex gel (CHCl3/MeOH/H2O 7:3:0 then 7:3:0.5) to yield 2
(10 mg, 88%) as a gum: Rf ) 0.6 (CHCl3/MeOH/H2O 8:2:0.5);
[R]20D -5.3 (c 1, CHCl3/MeOH 2:1); 1H NMR (CD3OD, 400 MHz)
δ (ppm) 0.51-0.61 (m, 1H), 0.74-0.82 (m, 30H), 0.90-1.70 (m,
110H), 1.80-1.84 (m, 1H), 3.26-3.95 (m, 42H), 4.25 (d, J ) 7.6
Hz, 4H); 13C NMR (CD3OD, 100 MHz) δ (ppm) 19.7, 19.9, 20.0,
22.8, 22.9, 24.7, 24.8, 25.1, 26.4, 28.3, 29.1, 29.8, 29.9, 29.99,
30.03, 30.1, 30.2, 30.3, 32.0, 33.1, 37.1, 37.2, 37.3, 37.6, 37.7, 37.8,
38.0, 39.2, 39.7, 40.5, 41.1, 61.6, 61.8, 69.3, 69.4, 69.5, 70.5, 71.6,
72.2, 73.6, 73.9, 75.1, 75.3, 76.0, 78.2, 80.4, 103.7, 104.1; HRMS
(FAB) calcd for C105H202O26 (M + Na)+ 1902.4382, found
1902.4383.
A mixture of 17a (1.77 g, 1.2 mmol, 1 equiv) and palladium
dihydroxide (200 mg, 5% w/w) in THF/EtOH (50 mL, 1:1) was
stirred at room temperature overnight under hydrogen atmosphere.
The reaction mixture was warmed (40-45 °C) and filtered on
Celite. The filtrate was concentrated under reduced pressure to yield
1a (1.2 g, 80%) as a colorless oil: Rf ) 0.3 (PE/EtOAc 8:2); [R]20
D
1
+8.4 (c 1, CHCl3); H NMR (CDCl3, 400 MHz) δ (ppm) 0.58-
0.66 (m, 1H), 0.84-0.89 (m, 30H), 1.05-1.83 (m, 110H), 1.86-
1.95 (m, 1H), 2.22-2.29 (m, 2H), 3.4-3.7 (m, 18H); 13C (CDCl3,
100 MHz) δ (ppm) 19.66, 19.73, 19.77, 22.7, 22.8, 24.4, 24.5, 24.9,
26.2, 28.0, 28.8, 29.5, 29.7, 29.8, 29.9, 30.0, 32.8, 36.8, 37.1, 37.2,
37.34, 37.39, 37.44, 37.5, 38.8, 39.4, 40.2, 40.8, 63.06, 63.08, 68.70,
70.9, 71.0, 71.9, 78.4; HRMS (ESI+) calcd for C81H162O6 (M +
Na)+ 1254.2269, found 1254.2264; (M + K)+ 1270.2009, found
1270.2015. Anal. Calcd for C81H162O6: C, 78.96; H, 13.25.
Found: C, 79.26; H, 13.25.
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methoxy]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-sn-diglycer-
ol 1b. Compound 1b was prepared from diol 9b (1.0 g, 2.0 mmol)
and alcohol 8 (2.78 g, 6.0 mmol) following a procedure similar to
that of 1a. Flash chromatography on silica gel (PE/EtOAc 99:1)
provided 17b (1.32 g, 45%) as a colorless oil: Rf ) 0.66 (PE/
EtOAc 90:10); [R]20D +1.7 (c 1.10, CHCl3); 1H NMR (CDCl3, 400
MHz) δ (ppm) 0.83-0.87 (m, 31H), 1.06-1.66 (m, 90H), 1.68-
1.78 (m, 2H), 1.91-1.99 (m, 1H), 2.13-2.24 (m, 2H), 3.28 (d, J
) 7.1 Hz, 4H), 3.39 (t, J ) 6.6 Hz, 4H), 3.43 (t, J ) 6.6 Hz, 8H),
3.50-3.70 (m, 10H), 4.54 (s, 4H), 7.26-7.35 (m, 10H); 13C NMR
(CDCl3, 100 MHz) δ (ppm) 19.7, 19.77, 19.84, 19.86, 22.7, 22.8,
24.5, 24.56, 24.58, 24.9, 26.2, 26.3, 29.61, 29.62, 29.73, 29.75,
29.82, 29.9, 32.86, 32.88, 34.0, 39.8, 69.0, 70.4, 70.8, 71.2, 71.8,
73.4, 75.7, 78.0, 127.6, 127.7, 128.4, 138.5. Anal. Calcd for
C91H166O8: C, 78.73; H, 12.05. Found: C, 78.49; H, 12.17.
Titled product 1b was obtained from 17b in 85% yield as a
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methylene]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1,1′-di-O-
(phosphatidylcholine)-sn-diglycerol 3. Nitrogen gas was bub-
bled in a solution of phosphoryl chloride (12 mL, 130 mmol, 1.7
equiv) in dry toluene (5 mL) for 15 min. 2-Bromoethanol (5.4
mL, 76 mmol, 1 equiv) was slowly added over 30 min, HCl was
trapped by an aqueous solution (2 M KOH), and the reaction
mixture was stirred at room temperature overnight under nitrogen
atmosphere. The residue was distilled under reduced pressure
(120 °C, 10 mmHg) to yield bromoethyldichlorophosphate (5.52
g).54
colorless oil: Rf ) 0.23 (PE/EtOAc 85:15); [R]20 +9.1 (c 1.00,
D
CHCl3); 1H NMR (CDCl3, 400 MHz) δ (ppm) 0.80-0.88 (m, 31H),
1.06-1.67 (m, 90H), 1.68-1.78 (m, 2H), 1.91-1.98 (m, 1H),
2.14-2.28 (m, 4H), 3.28 (d, J ) 7.1 Hz, 4H), 3.40 (t, J ) 6.6 Hz,
4H), 3.43 (t, J ) 6.6 Hz, 8H), 3.50-3.70 (m, 10H); 13C NMR
(CDCl3, 100 MHz) δ (ppm) 19.70, 19.71, 19.76, 19.83, 22.71,
22.74, 22.76, 22.80, 24.4, 24.55, 24.56, 24.9, 26.18, 26.25, 28.05,
28.9, 29.55, 29.60, 29.7, 29.8, 29.9, 32.85, 34.01, 37.1, 37.2, 37.36,
37.39, 37.42, 37.46, 37.48, 37.52, 37.6, 38.8, 39.4, 39.8, 63.2, 68.7,
71.0, 71.2, 71.9, 75.7, 78.4; HRMS (ESI+) calcd for C77H154O8
(M + Na)+ 1230.1541, found 1230.1524; (M + K)+ 1246.1281,
found 1246.1234. Anal. Calcd for C77H154O8: C, 76.56; H, 12.85.
Found: C, 76.33; H, 12.90.
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methylene]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1,1′-di-O-
[2,3,6-tri-O-acetyl-4-O-(2,3,4,6-tetra-O-acetyl-â-D-galactopyra-
nosyl)-â-D-glucopyranosyl]-sn-diglycerol 2. Glycosylation. To a
mixture of diol 1a (21 mg, 0.017 mmol, 1 equiv) and peracetylated
ethylthiolactoside (35 mg, 0.051 mmol, 3 equiv) in dichloromethane
(3 mL) were added N-iodosuccinimide (15.3 mg, 0.068 mmol, 4
equiv) and triethylsilyl triflate (1.5 µL, 0.007 mmol, 0.4 equiv) at
0 °C under nitrogen atmosphere. The reaction mixture was stirred
at room temperature for 10 min. Triethylamine then dichlo-
romethane were added, and the organic phase was washed with
10% aqueous Na2S2O3, water, and saturated NH4Cl. The combined
organic phases were dried (MgSO4) and concentrated under reduced
To a solution of bromoethyldichlorophosphate (118 mg, 0.49
mmol, 20 equiv) in dry dichloromethane (2 mL) were slowly added
triethylamine (136 µL, 0.97 mmol, 40 equiv) and a solution of diol
1a (30 mg, 0.024 mmol, 1 equiv) in dry dichloromethane (2 mL).
The reaction mixture was stirred at room temperature for 3 days
under darkness and nitrogen atmosphere, 2:1 CHCl3/MeOH and
brine were added, and the aqueous phase was washed with 2:1
CHCl3/MeOH. The combined organic phases were dried (MgSO4)
and concentrated under reduced pressure. The residue was purified
on Sephadex LH-20 gel (CHCl3/MeOH 2:1) and then on silica gel
(CHCl3/MeOH 9:1 then 8:2) to yield the bromide derivative (35
mg, 90%) as a colorless oil: Rf ) 0.3 (CHCl3/MeOH 2:1). Due to
its relative instability, the compound was not further characterized
and was used rapidly in the next step.
A solution of the previous bromide derivative (35 mg, 0.022
mmol, 1 equiv) in CHCl3/i-PrOH/H2O (2.7:2.25:0.75) and trim-
ethylamine (1.95 mL, 31-35% in EtOH) was stirred at room
temperature for 4 days. CHCl3/MeOH (2:1) and aqueous NaCl were
added, and the aqueous phase was extracted with 2:1 CHCl3/MeOH.
The combined organic phases were dried (MgSO4) and concentrated
under reduced pressure. The residue was purified on LH-20
Sephadex gel (CHCl3/MeOH 2:1) and then on silica gel (CHCl3/
(54) Chang Chung, Y.; Hong Chiu, Y.; Wei Wu, Y.; Tai Tao, Y.
Biomaterials 2005, 26, 2313-2324.
J. Org. Chem, Vol. 72, No. 22, 2007 8275

Brard et al.
MeOH/H2O/NH4OH 65:35:5.5:0.5) to yield 3 (9.5 mg, 25%) as a
gum: Rf ) 0.1 (CHCl3/MeOH/H2O/NH4OH 65:35:5.5:0.5); IR
Rf ) 0.8 (CH2Cl2/Et2O 98:2); 1H NMR (CDCl3, 400 MHz) δ (ppm)
0.62 (m, 1H), 0.80-0.88 (m, 30H), 1.01-1.80 (m, 110H), 1.90
(m, 1H), 3.28-3.65 (m, 18H); 13C NMR (CDCl3, 100 MHz) δ
(ppm) 19.6, 19.65, 19.68, 19.8, 22.6, 22.7, 24.3, 24.47, 24.49, 24.8,
26.1, 28.0, 28.8, 29.5, 29.6, 29.7, 30.0, 31.6, 32.8, 36.7, 37.0, 37.1,
37.3, 37.4, 37.4, 37.5, 38.8, 39.4, 40.2, 40.7, 52.1, 68.9, 70.1, 71.8,
77.88, 77.91; HRMS (ESI+) calcd for C81H160N6O4 (M + Na)+
1304.2399, found 1304.2454.
1
(neat) ν (cm-1) 1087, 1232, 1467-1377, 2340, 2921-2852; H
NMR (CD3OD/CDCl3 1:1, 400 MHz) δ (ppm) 0.61-0.63 (m, 1H),
0.83-0.87 (m, 30H), 1.06-1.75 (m, 110H), 1.90 (m, 1H), 3.32 (s,
18H), 3.40-3.65 (m, 14H), 3.73 (m, 4H), 3.90-3.93 (m, 4H), 4.24
(m, 4H); 13C NMR (CD3OD/CDCl3 1:1, 100 MHz) δ (ppm) 20.08,
20.13, 20.18, 20.19, 23.1, 23.2, 25.3, 25.6, 27.0, 28.8, 29.5, 30.4,
30.5, 30.56, 30.58, 32.5, 33.6, 37.6, 38.0, 38.1, 38.2, 40.2, 41.0,
41.5, 54.6, 60.2, 67.0, 69.6, 71.3, 72.5, 77.9; 31P NMR (CD3OD/
CDCl3 1:1, 162 MHz) δ (ppm) 0.64 (s, 1P).
Diamine 20a. A mixture of diazide 22a (90 mg, 0.07 mmol, 1
equiv) and palladium on activated carbon (9 mg, 10% w/w) in THF/
EtOH (10 mL, 1:1) was stirred at room temperature for 5 h under
hydrogen atmosphere. The reaction mixture was filtered on Celite,
and the solvents were evaporated under reduced pressure to yield
the diamine 20a (77 mg, 90%) as a yellow oil: Rf ) 0.5 (MeOH/
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methylene]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1′-O-(phos-
phatidylcholine)-sn-diglycerol 4. To a solution of diol 1a (50 mg,
0.04 mmol, 1 equiv) in benzene (2 mL) were added triethylamine
(113 µL, 80 mmol, 20 equiv) and 2-chloro-2-oxo-1,2,3-dioxaphos-
pholane (74 µL, 80 mmol, 20 equiv) at 0 °C, and the yellow mixture
was stirred at room temperature for 36 h. The precipitate was filtered
off, washed with toluene, and the solvents were evaporated under
reduced pressure. The residue in acetonitrile was placed in an
autoclave, and trimethylamine (1 mL) was added at -80 °C. In
the autoclave, the reaction mixture was then warmed at 65 °C for
48 h and was concentrated under reduced pressure. The residue
was purified on silica gel (CHCl3/MeOH/H2O/NH4OH 60:40:0:0
then 65:35:5.5:0.5) to yield 4 (26 mg, 46%) as a gum: Rf ) 0.6
1
Et3N 99:1); H NMR (CDCl3, 400 MHz) δ (ppm) 0.60-0.63 (m,
1H), 0.83-0.87 (m, 30H), 0.98-1.93 (m, 111H), 3.07-3.12 (m,
2H), 3.21-3.25 (m, 2H), 3.41-3.77 (m, 18H); 13C NMR (CDCl3,
100 MHz) δ (ppm) 19.5, 19.6, 19.66, 19.74, 22.6, 22.7, 24.40,
24.44, 24.5, 24.8, 26.0, 28.0, 28.59, 28.60, 29.59, 29.63, 29.68,
29.74, 29.81, 29.83, 29.9, 31.7, 32.80, 32.83, 33.0, 36.7, 36.9, 37.0,
37.3, 37.40, 37.44, 37.5, 39.4, 40.1, 40.5, 41.3, 68.8, 68.9, 70.4,
72.0, 73.9.
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methoxy]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1,1′-O-diamino-
sn-diglycerol 20b. Compound 20b was prepared from diol 1b (200
mg, 0.17 mmol, 1 equiv) following a procedure similar to that of 20a.
(CHCl3/MeOH/H2O/NH4OH 65:35:5.5:0.5); [R]20 -2.2 (c 1,
D
1
CHCl3/MeOH 2:1); H NMR (CD3OD/CDCl3 1:1, 400 MHz) δ
Dimesylate. The crude material was purified by flash chroma-
(ppm) 0.61-0.63 (m, 1H), 0.83-0.87 (m, 30H), 1.06-1.75 (m,
110H), 1.90 (m, 1H), 3.35 (s, 9H), 3.44-3.68 (m, 14H), 3.75 (m,
2H), 3.89-3.96 (m, 4H), 4.26 (m, 2H); 13C NMR (CD3OD/CDCl3
1:1, 100 MHz) δ (ppm) 20.0, 20.07, 20.14, 23.0, 23.1, 24.98, 25.02,
25.4, 25.8, 26.7, 28.5, 29.0, 30.1, 30.3, 30.4, 30.5, 32.2, 33.32,
33.34, 37.3, 37.65, 37.73, 37.8, 37.9, 38.0, 38.1, 38.2, 39.3, 39.9,
40.7, 41.3, 47.0, 51.9, 62.4, 62.5, 65.3, 65.4, 67.58, 67.64, 69.4,
69.6, 71.0, 71.1, 72.3, 78.0; 31P NMR (CD3OD/CDCl3 1:1, 162
MHz) δ (ppm) 1.69 (s, 1P); HRMS (FAB) calcd for C86H175NO9P
(M + H)+ 1397.3005, found 1397.3010.
tography on silica gel (PE/EtOAc 9:1) to yield the dimesylate
derivative (95%) as a colorless oil: Rf ) 0.5 (PE/EtOAc 8:2); H
1
NMR (CDCl3, 400 MHz) δ (ppm) 0.83-0.88 (m, 31H), 1.01-
1.63 (m, 90H), 1.68-1.77 (m, 2H), 1.95 (dt, J ) 12.7, 7.2 Hz,
1H), 2.12-2.23 (m, 2H), 2.97 (s, 6H), 3.28 (d, J ) 7.1 Hz, 4H),
3.39 (t, J ) 6.6 Hz, 4H), 3.43 (t, J ) 6.6 Hz, 4H), 3.46-3.69 (m,
10H), 4.18 (ddd, J ) 1.0, 5.7, 10.9 Hz, 2H), 4.31 (dd, J ) 3.6,
10.9 Hz, 2H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 19.5, 19.56,
19.58, 19.60, 19.63, 19.67, 19.73, 22.6, 22.7, 24.3, 24.45, 24.47,
24.8, 26.1, 26.16, 26.18, 28.0, 28.8, 29.46, 29.51, 29.57, 29.60,
29.7, 29.75, 29.77, 32.8, 33.9, 36.87, 36.89, 36.96, 36.97, 37.26,
37.30, 37.32, 37.35, 37.38, 37.40, 37.43, 37.47, 39.3, 39.7, 69.00,
69.04, 69.1, 69.70, 69.73, 71.1, 71.9, 75.6, 76.3.
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methoxy]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1,1′-O-diamino-
sn-diglycerol 20a. Dimesylate. To a mixture of diol 1a (200 mg,
0.16 mmol, 1 equiv) and triethylamine (67.9 µL, 0.49 mmol, 3
equiv) in dry dichloromethane (10 mL) was added mesyl chloride
(75.6 µL, 1.02 mmol, 6 equiv) at 0 °C, and the reaction mixture
was stirred overnight at room temperature. Water was added, and
the organic phase was washed with saturated NaHCO3 and brine.
The combined organic phases were dried (MgSO4) and concentrated
under reduced pressure. The residue was purified by flash chro-
matography on silica gel (PE/EtOAc 9:1) to yield the dimesylate
derivative (214 mg, 95%) as a colorless oil: Rf ) 0.5 (PE/EtOAc
Diazide 22b. The crude material was purified by flash chroma-
tography on silica gel (CH2Cl2/Et2O 98:2) to yield the diazide 22b
(90%) as a colorless oil: Rf ) 0.8 (CH2Cl2/Et2O 98:2); [R]20D +3.2
1
(c 1.0, CHCl3); H NMR (CDCl3, 400 MHz) δ (ppm) 0.83-0.87
(m, 31H), 1.06-1.66 (m, 90H), 1.68-1.78 (m, 2H), 1.95 (dt, J )
12.6, 7.5 Hz, 1H), 2.13-2.24 (m, 2H), 3.28 (d, J ) 7.1 Hz, 4H),
3.33 (m, 4H), 3.38 (t, J ) 6.6 Hz, 4H), 3.41-3.65 (m, 14H); 13C
NMR (CDCl3, 100 MHz) δ (ppm) 19.6, 19.65, 19.68, 19.8, 22.6,
22.7, 24.3, 24.46, 24.48, 24.8, 26.1, 26.18, 26.20, 28.0, 28.7, 28.8,
29.47, 29.52, 29.6, 29.7, 29.8, 32.77, 32.79, 33.9, 36.97, 36.99,
37.05, 37.08, 37.28, 37.34, 37.40, 37.44, 37.5, 39.4, 39.7, 52.1,
68.88, 68.90, 70.1, 71.0, 71.1, 71.8, 75.6, 77.85, 77.89. Anal. Calcd
for C77H152N6O6: C, 73.51; H, 12.18; N, 6.68. Found: C, 73.82;
H, 12.71; N, 6.63.
1
8:2); H NMR (CDCl3, 400 MHz) δ (ppm) 0.61 (m, 1H), 0.83-
0.88 (m, 30H), 1.02-1.81 (m, 110H), 1.88 (m, 1H), 3.04 (s, 6H),
3.41-3.69 (m, 14H), 4.24 (dd, J ) 5.7, 10.8 Hz, 2H), 4.38 (dd, J
) 3.6, 10.8 Hz, 2H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 19.5,
19.57, 19.58, 19.61, 19.64, 19.67, 19.74, 22.6, 22.7, 24.4, 24.46,
24.48, 24.8, 26.1, 28.0, 28.7, 28.8, 29.5, 29.57, 29.60, 29.63, 29.68,
29.71, 29.73, 29.8, 30.0, 31.6, 32.76, 32.78, 36.7, 36.8, 36.9, 37.0,
37.27, 37.31, 37.33, 37.35, 37.37, 37.39, 37.40, 37.44, 37.48, 37.50,
38.8, 39.3, 40.1, 40.7, 69.0, 69.1, 69.70, 69.73, 71.9, 76.4.
Diamine 20b. The crude material was filtered on Celite, and
the solvents were evaporated under reduced pressure to yield the
diamine 20b (90%) as a yellow oil: Rf ) 0.5 (MeOH/Et3N 99:1);
[R]20D +12.9 (c 0.91, CHCl3); 1H NMR (CDCl3, 400 MHz) δ (ppm)
0.82-0.86 (m, 31H), 1.06-1.66 (m, 96H), 1.91-1.99 (m, 1H),
2.12-2.22 (m, 2H), 3.04-3.09 (m, 2H), 3.19-3.62 (m, 22H),
3.72-3.80 (m, 2H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 19.6,
19.67, 19.74, 22.6, 22.7, 24.41, 24.43, 24.5, 24.8, 26.0, 26.15, 26.17,
28.0, 28.8, 29.45, 29.51, 29.54, 29.56, 29.6, 29.69, 29.71, 29.74,
29.81, 29.84, 32.8, 33.9, 36.85, 36.93, 37.29, 37.30, 37.39, 37.43,
37.47, 37.50, 37.52, 39.4, 39.7, 41.4, 68.7, 68.9, 70.9, 71.09, 71.13,
72.1, 73.7, 75.57, 75.59; HRMS (ESI+) calcd for C77H156N2O6 (M
+ H)+ 1206.2041; (M + Na)+ 1228.1861, found 1206.2041;
1228.1914.
Diazide 22a. A mixture of the previous dimesylate derivative
(75 mg, 0.054 mmol, 1 equiv), sodium azide (10.5 mg, 0.15 mmol,
3 equiv), and tetrabutylammonium bromide (8.7 mg, 0.25 mmol,
0.5 equiv) in dry DMF (1 mL) was placed under nitrogen
atmosphere. The resulting orange and heterogeneous mixture was
stirred at reflux for 1 h. Water was added, and the aqueous phase
was extracted with Et2O. The combined organic phases were dried
(MgSO4) and concentrated under reduced pressure. The residue was
purified by flash chromatography on silica gel (CH2Cl2/Et2O 98:2)
to yield the diazide 22a derivative (62 mg, 90%) as a colorless oil;
8276 J. Org. Chem., Vol. 72, No. 22, 2007

Synthesis of Archaeal Bipolar Lipid Analogues
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methylene]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1,1′-O-dibe-
taine-sn-diglycerol 5a. A mixture of diamine 20a (90 mg, 0.07
mmol, 1 equiv) and activated glycine betaine 21 (56 mg, 0.22 mmol,
3 equiv) in DMF/Et3N (11 mL, 10:1) was stirred at room
temperature overnight. The reaction mixture was concentrated under
reduced pressure, and the residue was purified by flash chroma-
tography on silica gel (CHCl3/acetone/MeOH/NH4OH 8:6:5:1) to
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methylene]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1-O-acetyl-sn-
diglycerol 19. A mixture of diol 1a (1.01 g, 0.821 mmol, 1 equiv),
acetic anhydride (293 mg, 2.9 mmol, 3.5 equiv), and sodium acetate
(101 mg, 1.23 mmol, 1.5 equiv) in dry dichloromethane (50 mL)
was stirred at reflux for 6.5 h. Water was added, and the aqueous
phase was extracted with dichloromethane. The combined organic
phases were dried (MgSO4) and concentrated under reduced
pressure. The residue was purified by flash chromatography on silica
yield 5a (84 mg, 80%) as a red gum: Rf ) 0.4 (CHCl3/acetone/
1
MeOH/NH4OH 8:6:5:1); [R]20 +13.5 (c 0.81, CHCl3); H NMR
gel (PE/EtOAc 8:2) to yield 19 (512 mg, 49%) as a colorless oil:
D
1
Rf ) 0.5 (PE/EtOAc 8:2); [R]20 +9.4° (c 1, CHCl3); H NMR
(CDCl3, 400 MHz) δ (ppm) 0.60-0.63 (m, 1H), 0.83-0.87 (m,
30H), 1.00-1.80 (m, 110H), 1.91 (m, 1H), 3.39-3.7 (m, 18H),
3.44 (s, 18H), 4.58 (m, 2H), 4.75 (m, 2H); 13C NMR (CDCl3, 100
MHz) δ (ppm) 20.0, 20.1, 20.3, 23.05, 23.15, 25.2, 26.1, 26.6, 28.4,
30.1, 30.2, 30.4, 32.1, 33.20, 33.22, 33.24, 37.2, 37.7, 37.8, 37.85,
37.89, 39.8, 40.6, 55.4, 65.7, 69.0, 69.1, 72.2, 73.6, 165.4; HRMS
(ESI+) calcd for C91H184N4O6 (M2+) 714.7108, found 714.7103.
D
(CDCl3, 400 MHz) δ (ppm) 0.60 (m, 1H), 0.84-0.89 (m, 30H),
1.05-1.83 (m, 110H), 1.86-1.95 (m, 1H), 2.09 (s, 3H), 2.17 (t, J
) 6.2 Hz, 1H), 3.40-3.78 (m, 16H), 4.11 (dd, J ) 5.9, 11.6 Hz,
1H), 4.23 (dd, J ) 4.1, 11.6 Hz, 1H); 13C NMR (CDCl3, 100 MHz)
δ (ppm) 19.5, 19.56, 19.62, 19.67, 19.74, 20.9, 22.6, 22.7, 24.3,
24.46, 24.48, 24.8, 26.1, 28.0, 28.8, 29.5, 29.6, 29.7, 29.8, 30.0,
31.6, 32.8, 33.0, 36.7, 36.8, 36.9, 37.0, 37.1, 37.23, 37.33, 37.38,
37.44, 37.5, 38.8, 39.4, 40.1, 40.7, 63.08, 63.10, 64.1, 68.6, 68.90,
68.91, 70.19, 70.21, 70.9, 71.8, 71.9, 76.5, 78.2, 78.3, 171.0; HRMS
(ESI+) calcd for C83H164O7 (M + Na)+ 1296.2375, found 1296.2378.
Benzyloxymethyl Trisallyloxymethylmethane 25. Monoben-
zylation. To a solution of pentaerythritol 24 (20 g, 147 mmol, 1
equiv) and tetrabutylammonium iodide (1.5 g) in DMF (100 mL)
was added slowly at 0 °C sodium hydride (1.47 g, 36.8 mmol, 0.25
equiv). The reaction mixture was stirred at room temperature for 1
h. Benzyl bromide (4.4 mL, 36.8 mmol, 0.25 equiv) was added,
and the reaction mixture was stirred at room temperature for 12 h.
MeOH was added, and the mixture was concentrated under reduced
pressure. The residue was dissolved in EtOAc and washed with
water. The organic phase was concentrated under reduced pressure.
The residue was dissolved in dichloromethane, washed with water,
and extracted with EtOAc. The combined organic phases were dried
(MgSO4) and concentrated under reduced pressure to yield monoben-
zylated derivative (5.8 g, 72%) as a white solid: Rf ) 0.3 (EtOAc/
PE/MeOH 10:5:1); mp 73 °C; 1H NMR (CDCl3, 400 MHz) δ (ppm)
3.22 (br s, 3H), 3.48 (s, 2H), 3.68 (s, 6H), 4.48 (s, 2H), 7.29-7.32
(m, 5H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 45.02, 64.22, 72.23,
73.71, 127.53, 127.82, 128.50, 137.66. These analytical data are
in total agreement with previously published data.55
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methoxy]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1,1′-O-dibe-
taine-sn-diglycerol 5b. Compound 5b was prepared from 20b
following a similar procedure to that of 5a. The crude material
was purified by flash chromatography on silica gel (CHCl3/acetone/
MeOH/NH4OH 8:6:5:1) to yield 5b (80%) as a red gum: Rf ) 0.4
(CHCl3/acetone/MeOH/NH4OH 8:6:5:1); [R]20 +13.1 (c 0.81,
D
1
CHCl3); H NMR (CD3OD, 400 MHz) δ (ppm) 1.05-1.08 (m,
30H), 1.23-2.15 (m, 96H), 3.46-3.83 (m, 26H), 3.54 (s, 18H),
4.39 (s, 4H); 13C NMR (CD3OD, 100 MHz) δ (ppm) 20.65, 20.71,
20.86, 20.91, 23.7, 23.8, 26.0, 26.4, 27.0, 27.8, 29.6, 30.1, 30.3,
31.1, 31.2, 31.27, 31.33, 31.52, 31.57, 31.61, 34.39, 34.42, 35.3,
38.8, 38.9, 38.98, 39.04, 41.0, 40.5, 41.8, 55.3, 66.3, 69.8, 70.1,
72.5, 72.7, 73.1, 77.0, 78.7, 78.9, 164.9; HRMS (ESI+) calcd for
C87H176N4O8 (M2+) 702.6744, found 702.6745.
Monocationic Tetraether 23. Compound 23 was prepared from
22a following a similar procedure to that of 5a except that the
reduction of 22a was done in 1:1 CHCl3/MeOH instead of 1:1 THF/
MeOH: 1H NMR (CDCl3, 400 MHz) δ (ppm) 0.57-0.65 (m, 1H),
0.83-0.88 (m, 30H), 1.04-1.97 (m, 111H), 3.35-3.70 (m, 18H),
3.46 (s, 9H), 4.60 (d, J ) 14.4 Hz, 1H), 4.71 (d, J ) 14.4 Hz, 1H);
13C NMR (CDCl3, 100 MHz) δ (ppm) 19.56, 19.63, 19.7, 22.6,
22.7, 24.28, 24.34, 24.41, 24.44, 24.8, 26.0, 26.1, 27.9, 28.7, 29.52,
29.54, 29.57, 29.63, 29.65, 29.69, 29.77, 29.84, 29.93, 29.94, 31.6,
32.71, 32.73, 32.8, 33.0, 36.7, 36.86, 36.93, 37.2, 37.35, 37.42,
37.5, 38.7, 39.3, 40.1, 40.31, 40.33, 40.7, 43.90, 43.92, 45.8, 54.7,
65.4, 68.5, 68.6, 68.8, 69.92, 69.93, 70.86, 70.89, 71.71, 76.74,
76.8, 78.2, 78.3, 163.0; HRMS (ESI+) calcd for C86H172N2O5Cl
(M+) 1348.2949, found 1348.2907.
3,3′-O-[1,33-Tetratriaconta-(cis-16,19-methylidene)methylene]-
2,2′-di-O-[3,7-(R),11-(R),15-tetramethylhexadecyl]-1-O-benzyl-
sn-diglycerol 18. To a solution of diol 1a (584 mg, 0.47 mmol, 1
equiv) and [15]-crown-5 ether (50 µL) in dry THF (18 mL) was
added sodium hydride (60% in mineral oil, 18.8 mg, 0.47 mmol, 1
equiv) at 0 °C, and the reaction mixture was stirred at 0 °C for 10
min. Benzyl bromide (84 µL, 0.70 mmol, 1.5 equiv) was added
dropwise, and the reaction mixture was stirred under reflux for 36
h. Et2O and water were added, and the aqueous phase was extracted
with Et2O and EtOAc. The combined organic phases were dried
(MgSO4) and concentrated under reduced pressure. The residue was
purified by flash chromatography on silica gel (PE/EtOAc 9:1 then
Triallylation. To a solution of the previous triol (1.45 g, 6.4
mmol, 1 equiv) and allyl bromide (2.8 mL, 32 mmol, 5 equiv) in
DMF (90 mL) was added slowly at 0 °C sodium hydride (60% in
mineral oil, 1.02 g, 25.6 mmol, 4 equiv), and the reaction mixture
was stirred at room temperature for 12 h. MeOH was added, and
the reaction mixture was concentrated under reduced pressure. The
residue was purified by flash chromatography on silica gel (PE/
EtOAc 7:3) to yield 25 (1.80 g, 81%) as a colorless oil: Rf ) 0.76
1
(PE/EtOAc 6:4); H NMR (CDCl3, 400 MHz) δ (ppm) 3.56 (s,
6H), 3.60 (s, 2H), 4.02 (dt, J ) 5.4, 1.5 Hz, 6H), 4.58 (s, 2H), 5.20
(dq, J ) 10.4, 1.5 Hz, 3H), 5.32 (dq, J ) 17.2, 1.5 Hz, 3H), 5.95
(ddt, J ) 10.4, 17.2, 5.4 Hz, 3H), 7.31-7.42 (m, 5H); 13C NMR
(CDCl3, 100 MHz) δ (ppm) 45.4, 69.3, 69.5, 72.2, 73.2, 116.1,
127.2 (2C), 128.2, 135.2, 139.0; HRMS (ESI+) calcd for C21H30O4
(M + Na)+ 369.2042, found 369.2036.
Benzyloxymethyl Tris(propanol-3-oxymethyl)methane 26. To
a solution of 25 (797 mg, 2.3 mmol, 1 equiv) in dry dioxane (20
mL) was added at 0 °C 9-borabicylco[3.3.1]nonane (41 mL, 20.7
mmol, 9 equiv), and the reaction mixture was stirred at room
temperature for 24 h. An aqueous solution of sodium hydroxide
(50 mL, 3M) and a solution of H2O2 (8 mL, 30%) were added at
0 °C, and the resulting mixture was stirred at room temperature
for 12 h. The reaction mixture was extracted with EtOAc, and the
combined organic phases were dried (MgSO4) and concentrated
under reduced pressure. The residue was purified by flash chro-
matography on silica gel (EtOAc/PE/MeOH 10:5:1) to yield 26
7:3) to yield 18 (320 mg, 51%) as a yellow oil: Rf ) 0.5 (PE/
1
EtOAc 9:1); [R]20 +3.3 (c 1.04, CHCl3); H NMR (CDCl3, 400
D
MHz) δ (ppm) 0.60 (m, 1H), 0.75-0.80 (m, 30H), 1.00-1.80 (m,
110H), 1.80-1.96 (m, 1H), 2.17 (t, J ) 6.2 Hz, 1H), 3.34-3.66
(m, 18H), 4.48 (s, 2H), 7.18-7.26 (m, 5H); 13C NMR (CDCl3,
100 MHz) δ (ppm) 19.6, 19.67, 19.74, 22.6, 22.7, 24.4, 24.5, 24.8,
26.1, 28.0, 28.8, 29.47, 29.50, 29.65, 29.71, 29.8, 30.0, 31.6, 32.8,
36.7, 37.28, 37.34, 37.38, 37.44, 38.8, 39.4, 40.1, 40.7, 63.1, 68.6,
68.9, 70.3, 70.7, 70.9, 71.7, 71.9, 73.3, 77.9, 78.2, 127.5, 127.6,
128.3, 138.4.
(55) Dunn, T. J.; Neumann, W. L.; Rogic, M. M.; Woulfe, S. R. J. Org.
Chem. 1990, 55, 6368-6373.
J. Org. Chem, Vol. 72, No. 22, 2007 8277

Brard et al.
(830 mg, 90%) as a gum: Rf ) 0.25 (EtOAc/PE/MeOH 10:5:1);
1H NMR (CDCl3, 400 MHz) δ (ppm) 1.77 (m, 6H), 3.21 (br s,
3H), 3.40 (s, 2H), 3.41 (s, 6H), 3.56 (t, J ) 4.8 Hz, 6H), 3.72 (t,
J ) 5.4 Hz, 6H), 4.47 (s, 2H), 7.27-7.36 (m, 5H); 13C NMR
(CDCl3, 100 MHz) δ (ppm) 31.6, 44.7, 61.5, 69.6, 70.5, 70.9, 73.3,
127.4, 127.5, 128.3, 138.4. Anal. Calcd for C21H36O7: C, 62.98;
H, 9.06. Found: C, 62.65; H, 8.75.
7.18 (m, 6H), 7.24-7.35 (m, 21H), 7.40 (t, J ) 7.5 Hz, 3H), 7.49
(m, 6H), 7.75 (d, J ) 8.4 Hz, 6H), 7.87 (d, J ) 8.5 Hz, 6H), 7.96
(d, J ) 8.4 Hz, 6H), 8.02 (d, J ) 8.4 Hz, 6H); 13C NMR (CDCl3,
100 MHz) δ (ppm) 30.1, 45.4, 63.2, 66.0, 66.1, 67.3, 68.6, 69.2,
70.6, 70.9, 71.9, 98.1, 128.7, 128.8, 128.9, 129.0, 129.4, 129.5,
129.8, 130.1, 130.2, 130.25, 130.29, 133.5, 133.6, 133.8, 165.8,
165.89, 166.6; HRMS (ESI+) calcd for C116H108O34 (M + Na)+
2067.6618, found 2067.6662. Anal. Calcd for C116H108O34: C,
68.09; H, 5.32. Found: C, 68.37; H, 5.50.
Benzyloxymethyl Tris(1-(perbenzoyl-D-mannosyl)propan-3-
oxymethyl)methane 28a. To a mixture of the mannosyl donor 27a
(2.5 g, 3.4 mmol, 20 equiv) and triol 26 (68 mg, 0.17 mmol, 1
equiv) in dry dichloromethane (5 mL) was added a solution of
trimethylsilyl trifluoromethane sulfonate (62 µL, 5% in dichlo-
romethane), and the reaction mixture was stirred at room temper-
ature for 12 h. NaHCO3 (1 g) was added, and the reaction mixture
was concentrated under reduced pressure. The residue was purified
by flash chromatography on silica gel (PE/EtOAc 6:4) to yield 28a
Tris(1-(perbenzoyl lactosyl)propan-3-oxymethyl)methanol 29b.
A mixture of 28b (120 mg, 0.04 mmol, 1 equiv) and palladium on
activated carbon (24 mg, 20% w/w) in CHCl3/EtOH (20 mL, 1:1)
was stirred at room temperature for 3 h under hydrogen atmosphere.
The reaction mixture was filtered on Celite, and the filtrate was
concentrated under reduced pressure. The residue was purified by
flash chromatography on silica gel (PE/EtOAc 1:1) to yield 29b
(327 mg, 90%) as a white solid: Rf ) 0.23 (PE/EtOAc 6:4); [R]20
(105 mg, 90%) as a white solid: Rf ) 0.5 (PE/EtOAc 1:1); [R]20
D
D
1
1
-25.0 (c 1.5, CHCl3); H NMR (CDCl3, 400 MHz) δ (ppm) 1.95
+53.1 (c 1.6, CHCl3); H NMR (CDCl3, 400 MHz) δ (ppm) 1.52
(m, 6H), 3.48 (s, 6H), 3.49 (s, 2H), 3.54 (m, 6H), 3.65 (m, 3H),
3.88 (m, 3H), 4.39-4.44 (m, 6H), 4.46 (m, 2H), 4.68 (dd, J )
12.0, 2.3 Hz, 3H), 5.06 (d, J ) 1.6 Hz, 3H), 5.69 (dd, J ) 3.0, 1.8
Hz, 3H), 5.91 (dd, J ) 7.4, 4.0 Hz, 3H), 6.12 (t, J ) 10.2 Hz, 3H),
7.17-7.43 (m, 32H), 7.47 (t, J ) 7.4 Hz, 3H), 7.56 (m, 6H), 7.82
(d, J ) 8.4 Hz, 6H), 7.93 (d, J ) 8.4 Hz, 6H), 8.03 (d, J ) 8.4 Hz,
6H), 8.09 (d, J ) 8.4 Hz, 6H); 13C NMR (CDCl3, 100 MHz) δ
(ppm) 30.1, 45.9, 63.2, 66.3, 67.3, 68.3, 69.2, 70.2, 70.6, 70.9, 73.7,
98.1, 127.67, 127.72, 128.6, 128.7, 128.8, 128.9, 129.0, 129.4,
129.5, 129.8, 130.1, 130.19, 130.23, 130.3, 133.4, 133.5, 133.8,
139.3, 165.7, 165.9, 166.5; HRMS (ESI+) calcd for C123H114O34
(M + Na)+ 2157.7090, found 2157.7075.
(m, 6H), 2.79 (d, J ) 9.2 Hz, 3H), 2.87 (d, J ) 9.2 Hz, 3H), 3.02
(t, J ) 6.4 Hz, 6H), 3.22 (s, 2H), 3.30-3.38 (m, 3H), 3.58-3.67
(m, 7H), 3.70-3.75 (m, 6H), 3.80 (t, J ) 6.8 Hz, 3H), 4.15 (t, J )
9.4 Hz, 3H), 4.40 (dd, J ) 4.1, 12.2 Hz, 3H), 4.49 (d, J ) 10.8
Hz, 3H), 4.57 (d, J ) 7.9 Hz, 3H), 4.78 (d, J ) 7.9 Hz, 3H), 5.25-
5.37 (m, 6H), 5.61-5.71 (m, 9H), 7.06 (t, J ) 7.7 Hz, 6H), 7.13
(t, J ) 7.7 Hz, 6H), 7.20-7.50 (m, 48H), 7.54 (t, J ) 7.5 Hz, 3H),
7.65 (d, J ) 7.2 Hz, 6H), 7.81-7.84 (m, 12H), 7.86-7.94 (m,
24H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 29.6, 44.3, 61.1, 62.4,
65.6, 67.2, 67.5, 67.8, 69.9, 71.1, 71.4, 71.8, 72.9, 73.0, 76.1, 101.0,
101.2, 128.3, 128.4, 128.56, 128.60, 128.63, 128.7, 128.9, 129.3,
129.4, 129.5, 129.6, 129.67, 129.70, 129.8, 130.0, 133.2, 133.26,
133.30, 133.5, 133.6, 164.8, 165.2, 165.3, 165.5 (2C), 165.6, 165.9;
HRMS (ESI+) calcd for C197H174O58 (M + Na)+ 3490.0564, found
3490.0558; (M + K)+ calcd 3506.0303, found 3506.0339. Anal.
Calcd for C197H174O58: C, 68.20; H, 5.06. Found: C, 67.92; H,
5.08.
Benzyloxymethyl Tris(1-(perbenzoyl-D-lactosyl)propan-3-
oxymethyl)methane 28b. To a mixture of the lactosyl donor 27b
(3.9 g, 3.2 mmol, 20 equiv) and triol 26 (64 mg, 0.16 mmol, 1
equiv) in dry dichloromethane (25 mL) was added a solution of
trimethylsilyl trifluoromethane sulfonate (25 µL, 5% in dichlo-
romethane). The reaction mixture was stirred at room temperature
for 12 h. NaHCO3 (1 g) was added, and the reaction mixture was
concentrated under reduced pressure. The residue was purified by
flash chromatography on silica gel (PE/EtOAc 6:4) to yield 28b
Trimannosyl Cluster Tetraether 6. Preparation of 30a. A
solution of 29a (50 mg, 0.024 mmol, 1 equiv) in dry acetonitrile
was concentrated under reduced pressure and dried under reduced
pressure for 2 h. The residue (29a) was dissolved in degassed dry
acetonitrile (0.5 mL), then dimethyl sulfide (16 µL, 192 mmol, 8
equiv) and benzoyl peroxide (26 mg, 96 mmol, 4 equiv) were added
at 0 °C. The reaction mixture was stirred at 0 °C for 3 h, EtOAc
was added, and the organic phase was washed with brine. The
combined organic phases were dried (MgSO4) and concentrated
under reduced pressure. The residue was purified by flash chro-
matography on silica gel (CH2Cl2/acetone 98:2) to yield 30a (33
mg, 80%) as a white solid. The product was rapidly characterized
(some impurities were found in the NMR spectra) and was used as
(541 mg, 95%) as a white solid: Rf ) 0.14 (PE/EtOAc 6:4); [R]20
D
1
+34.5 (c 1.1, CHCl3); H NMR (CDCl3, 400 MHz) δ (ppm) 1.52
(m, 6H), 2.86 (dd, J ) 8.9, 12.7 Hz, 6H), 3.02 (t, J ) 6.1 Hz, 6H),
3.06 (s, 2H), 3.30-3.38 (dt, J ) 6.3, 13.2 Hz, 3H), 3.55-3.68 (m,
9H), 3.70 (dt, J ) 6.3, 13.2 Hz, 3H), 3.79 (m, 3H), 4.15 (t, J ) 9.4
Hz, 3H), 4.23 (s, 2H), 4.35-4.51 (dd, J ) 4.1, 12.2 Hz, 6H), 4.49
(d, J ) 7.9 Hz, 3H), 4.78 (d, J ) 7.9 Hz, 3H), 5.25-5.37 (m, 6H),
5.61-5.71 (m, 9H), 7.06 (t, J ) 7.8 Hz, 6H), 7.13 (t, J ) 7.6 Hz,
8H), 7.17-7.56 (m, 54H), 7.65 (d, J ) 8.4 Hz, 6H), 7.82 (d, J )
8.1 Hz, 12H), 7.91 (m, 24H); 13C NMR (CDCl3, 100 MHz) δ (ppm)
29.7, 45.0, 61.1, 62.4, 67.48, 67.53, 69.4, 69.9, 71.4, 71.78, 71.83,
72.9, 73.0, 76.0, 101.0, 101.3, 127.0, 127.4, 128.3, 128.4, 128.61,
128.64, 128.7, 128.9, 129.3, 129.4, 129.5, 129.6, 129.68, 129.74,
129.78, 129.82, 130.0, 133.2, 133.3, 133.5, 133.6, 139.0, 164.8,
165.2, 165.3, 165.4, 165.6, 165.9; HRMS (ESI+) calcd for
1
such in the next step: Rf ) 0.3 (PE/EtOAc 6:4); H (CDCl3, 400
MHz) δ (ppm) 1.94 (m, 6H), 2.1 (s, 3H), 3.38 (s, 6H), 3.51 (m,
8H), 3.62 (m, 3H), 3.86 (m, 3H), 4.35-4.44 (m, 6H), 4.52 (s, 2H),
4.66 (dd, J ) 12.0, 2.3 Hz, 3H), 5.04 (d, J ) 1.6 Hz, 3H), 5.66
(dd, J ) 3.1, 1.8 Hz, 3H), 5.87 (dd, J ) 7.4, 4.0 Hz, 3H), 6.09 (t,
J ) 10.2 Hz, 3H), 7.15-7.55 (m, 36H), 7.75 (d, J ) 8.4 Hz, 6H),
7.87 (d, J ) 8.4 Hz, 6H), 7.97 (d, J ) 8.4 Hz, 6H), 8.02 (d, J )
8.4 Hz, 6H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 13.9, 29.8,
45.4, 62.9, 65.9, 67.0, 68.0, 68.9, 69.9, 70.2, 70.6, 75.8, 97.8, 128.4,
128.48, 128.52, 128.6, 129.1, 129.2, 129.4, 129.8, 129.87, 129.91,
130.0, 130.3, 133.1, 133.2, 133.5, 165.43, 165.44, 166.2.
C
204H180O58 (M + Na)+ 3580.1033, found 3580.1019.
Tris(1-(perbenzoyl-D-mannosyl)propan-3-oxymethyl)metha-
nol 29a. A mixture of 28a (80 mg, 0.04 mmol, 1 equiv) and
palladium on activated carbon (15 mg, 20% w/w) in CHCl3/MeOH
(6 mL, 1:1) was stirred at room temperature for 3 h under hydrogen
atmosphere. The reaction mixture was filtered on Celite, and the
filtrate was concentrated under reduced pressure. The residue was
purified by flash chromatography on silica gel (PE/EtOAc 6:4) to
yield 29a (69 mg, 91%) as a white solid: Rf ) 0.5 (PE/EtOAc
6:4); [R]20D -34.0 (c 1.6, CHCl3); 1H NMR (CDCl3, 400 MHz) δ
(ppm) 1.90 (m, 6H), 2.90 (t, J ) 5.9 Hz, 1H), 3.46 (s, 6H), 3.51 (t,
J ) 6.1 Hz, 6H), 3.59 (m, 3H), 3.75 (d, J ) 5.0 Hz, 2H), 3.88 (m,
3H), 4.35 (m, 3H), 4.41 (dd, J ) 12.0, 4.4 Hz, 3H), 4.63 (dd, J )
12.0, 2.3 Hz, 3H), 5.02 (d, J ) 1.6 Hz, 3H), 5.62 (dd, J ) 3.0, 1.8
Hz, 3H), 5.84 (dd, J ) 7.4, 4.0 Hz, 3H), 6.05 (t, J ) 10.0 Hz, 3H),
Preparation of 6 (Protected). A mixture of 30a (20 mg, 0.010
mmol, 1 equiv) and 19 (14 mg, 0.012 mmol, 1.2 equiv) in dry
dichloromethane (2 mL) was concentrated under reduced pressure
and dried under reduced pressure for 2 h. The residue was dissolved
in CH2Cl2/THF (0.3 mL, 1:1), molecular sieves (15 mg, 4 Å) was
added, and the reaction mixture was stirred at room temperature
for 15 min. A solution of N-iodosuccinimide (1.25 equiv) and triflic
acid (0.12 equiv) in CH2Cl2/THF (0.4 mL, 1:1) was added at 0 °C,
and the reaction mixture was stirred for 5 min. Pyridine (0.5 mL)
and dichloromethane were added, and the organic phase was washed
8278 J. Org. Chem., Vol. 72, No. 22, 2007

Synthesis of Archaeal Bipolar Lipid Analogues
with aqueous 1 M Na2S2O3 and 10% aqueous NaHCO3. The
combined organic phases were dried (MgSO4) and concentrated
under reduced pressure. The residue was purified by flash chro-
matography on silica gel (PE/EtOAc 7:3) to yield 6 (protected)
71.8, 72.2, 73.3, 76.4, 101.4, 101.6, 128.7, 128.8, 128.9, 128.97,
129.03, 129.2, 129.7, 129.8, 129.9, 130.0, 130.05, 130.08, 130.11,
130.2, 130.4, 130.6, 133.6, 133.7, 133.8, 134.0, 134.1, 165.2,
165.55, 165.64, 165.8, 166.0, 166.2.
(19 mg, 60%, two steps) as a colorless oil: Rf ) 0.5 (PE/EtOAc
Preparation of 7 (Protected). Compound 7 (Protected) (28.5
mg, 60%, two steps) was prepared from 30b (40 mg, 0.010 mmol,
1 equiv) and 19 (14 mg, 0.0125 mmol, 1.2 equiv) following a
similar procedure to that of 7 (protected). Colorless oil: Rf ) 0.6
(PE/EtOAc 65:35); [R]20D +41.3 (c 1.1, CHCl3); 1H NMR (CDCl3,
400 MHz) δ (ppm) 0.62 (m, 1H), 0.81-0.88 (m, 30H), 1.02-1.93
(m, 111H), 2.07 (s, 3H), 2.89 (m, 6H), 3.07 (t, J ) 6.4 Hz, 6H),
3.17 (s, 2H), 3.34-3.81 (m, 37H), 3.86 (t, J ) 6.6 Hz, 3H), 4.10
(dd, J ) 5.8, 11.6 Hz, 1H), 4.21 (dd, J ) 4.0, 7.4 Hz, 1H), 4.25 (t,
J ) 9.4 Hz, 3H), 4.45 (s, 2H), 4.48 (dd, J ) 4.1, 12.2 Hz, 3H),
4.56 (d, J ) 10.6 Hz, 3H), 4.62 (d, J ) 7.9 Hz, 3H), 4.86 (d, J )
7.9 Hz, 3H), 5.35 (dd, J ) 3.4, 10.3 Hz, 3H), 5.35 (dd, J ) 8.0,
9.9 Hz, 3H), 5.71 (m, 6H), 5.77 (t, J ) 9.4 Hz, 3H), 7.14 (t, J )
7.7 Hz, 6H), 7.21 (t, J ) 7.7 Hz, 6H), 7.27-7.64 (m, 51H), 7.72
(d, J ) 8.5 Hz, 6H), 7.87-7.90 (m, 12H), 7.96-8.01 (m, 24H);
13C NMR (CDCl3, 100 MHz) δ (ppm) 19.6, 19.67, 19.74, 20.9,
22.6, 22.7, 24.3, 24.5, 24.8, 25.5, 26.08, 26.13, 28.0, 28.8, 28.9,
29.49, 29.52, 29.56, 29.62, 29.69, 29.72, 29.77, 29.82, 29.84, 29.99,
30.01, 31.6, 32.78, 32.84, 36.7, 37.28, 37.34, 37.4, 37.6, 38.6, 39.4,
40.1, 40.8, 45.6, 61.0, 61.7, 62.4, 67.5, 67.7, 68.9, 69.8, 69.9, 70.2,
71.3, 71.7, 71.8, 72.9, 76.0, 76.5, 96.0, 101.0, 101.2, 128.2, 128.3,
128.5, 128.55, 128.61, 128.64, 128.8, 129.3, 129.4, 129.47, 129.54,
129.6, 129.65, 129.74, 129.8, 130.0, 133.1, 133.2, 133.4, 133.5,
164.8, 165.1, 165.2, 165.37, 165.38, 165.5, 165.8; HRMS (ESI+)
calcd for C281H338O65Na (M + Na)+ 4775.3041, found 4775.3036.
1
7:3); [R]20 -24.3 (c 0.95, CHCl3); H NMR (CDCl3, 400 MHz)
D
δ (ppm) 0.57 (m, 1H), 0.80-0.88 (m, 30H), 0.98-1.80 (m, 110H),
1.94 (m, 7H), 2.1 (s, 3H), 3.38-3.70 (m, 33H), 3.86 (m, 3H), 4.05
(dd, J ) 5.8, 11.5 Hz, 1H), 4.18 (dd, J ) 4.1, 11.5 Hz, 1H), 4.36-
4.46 (m, 6H), 4.65-4.68 (m, 5H), 5.04 (d, J ) 1.6 Hz, 3H), 5.66
(dd, J ) 1.7, 3.0 Hz, 3H), 5.87 (dd, J ) 4.0, 7.4 Hz, 3H), 6.09 (t,
J ) 10.2 Hz, 3H), 7.19 (t, J ) 7.7 Hz, 6H), 7.25-7.37 (m, 21H),
7.41 (t, J ) 7.4 Hz, 3H), 7.51 (t, J ) 7.5 Hz, 6H), 7.78 (d, J ) 8.4
Hz, 6H), 7.89 (d, J ) 8.4 Hz, 6H), 7.99 (d, J ) 8.4 Hz, 6H), 8.05
(d, J ) 8.4 Hz, 6H); 13C NMR (CDCl3, 100 MHz) δ (ppm) 19.6,
19.67, 19.74, 19.8, 21.0, 22.7, 22.8, 24.4-24.6, 24.9, 26.16, 26.22,
28.0, 28.8, 29.6, 29.7-29.9, 30.1, 31.7, 32.9, 36.8, 36.9, 37.0, 37.1,
37.15, 37.24, 37.4, 37.4-37.6, 37.7, 38.8, 39.4, 40.2, 40.8, 45.2,
62.8, 64.2, 65.9, 66.9, 67.3, 67.6, 68.1, 68.8, 69.0, 69.9, 70.2-
70.3, 70.6, 71.1, 71.7, 71.8, 76.6, 77.9, 78.0, 96.3, 97.8, 128.3,
128.46, 128.50, 128.6, 129.0, 129.2, 129.4, 129.8, 129.85, 129.89,
130.0, 133.1, 133.2, 133.4, 165.4, 165.49, 165.51, 166.2; HRMS
(ESI+) calcd for C200H272O41 (M + Na)+ 3352.9297, found
3352.9113; (M + K)+ calcd 3368.8836, found 3368.8761.
Preparation of 6. To a solution of previous 6 (protected) (100
mg, 0.030 mmol, 1 equiv) in CH2Cl2/MeOH (20 mL, 1:1) was
added a 0.1 M solution of MeONa (5 mL), and the reaction mixture
was stirred at room temperature for 48 h. Amberlite resin (IR120)
was added, the reaction mixture was filtered, and the solvents were
evaporated under reduced pressure. The residue was purified on
LH-20 Sephadex gel (CH2Cl2/MeOH 1:1) to yield 7 (60 mg,
Preparation of 7. Compound 7 was prepared from 7 (protected)
following a similar procedure to that of 6. Compound 7 was found
to be insoluble in most of common organic solvents. Only 1H NMR
could be recorded in d5-pyridine: 1H NMR (d5-pyridine, 400 MHz)
δ (ppm) 0.70 (m, 1H), 0.88-0.97 (m, 30H), 1.10-2.02 (m, 117H),
3.52-4.54 (m, 74H), 4.78 (d, J ) 7.8 Hz, 3H), 4.88 (s, 2H), 5.09
(d, J ) 7.9 Hz, 3H); HRMS (ESI+) calcd for C132H252O43 (M +
Na)+ 2548.7430, found 2548.7441.
quantitative) as a colorless oil: Rf ) 0.5 (EtOAc/MeOH/H2O 5:3:
1
2); [R]20 -14.9 (c 1.05, CHCl3); H NMR (CD3OD/CDCl3 1:1,
D
400 MHz) δ (ppm) 0.61 (m, 1H), 0.83-0.88 (m, 30H), 1.04-1.89
(m, 117H), 3.33-3.83 (m, 56H), 4.64 (s, 2H), 4.75 (d, J ) 1.6 Hz,
3H); 13C NMR (CD3OD/CDCl3 1:1, 100 MHz) δ (ppm) 19.2-
19.3, 22.15, 22.24, 24.1, 24.2, 24.5, 25.3, 25.80, 25.83, 27.7, 28.4,
29.2-29.6, 31.3, 32.5, 36.5, 36.8-37.2, 39.1, 39.9, 49.2, 61.2, 61.3,
64.1, 64.3, 66.8, 66.9, 67.0-67.2, 67.8, 68.0, 68.5, 68.6, 69.4, 70.4-
70.6, 71.1, 71.4, 71.5, 72.3, 78.8, 78.9, 95.9, 99.9. HRMS (ESI+)
calcd for C114H222O28 (M + Na)+ 2062.5845, found 2062.5853.
Trilactosyl Cluster Tetraether 7. Preparation of 30b. Com-
pound 30b was prepared from 29b (100 mg, 0.029 mmol, 1 equiv)
following a similar procedure to that of 30a. The product was
rapidly characterized (some impurities were found in the NMR
spectra) and was used as such in the next step: Rf ) 0.1 (PE/
Acknowledgment. We are grateful to the Re´gion Bretagne
for grants to G.R. and C.N., and to the MENRT for a grant to
M.B. This work was also supported (grant to C.L.) by ‘‘Vaincre
La Mucoviscidose” (Paris, France). We are thankful to S. Pilard
(Universite´ de Picardie Jules Verne) and to W. Richter (Uni-
versita¨tsklinikum Jena) for recording, respectively, mass spectra
and FFEM.
1
EtOAc 6:4); H NMR (CDCl3, 400 MHz) δ (ppm) 1.52 (m, 6H),
2.1 (s, 3H), 2.86 (s, 6H), 3.02 (t, J ) 6.4 Hz, 6H), 3.06 (s, 2H),
3.30-3.38 (dt, J ) 6.3, 13.2 Hz, 3H), 3.55-3.68 (m, 9H), 3.70
(dt, J ) 6.3, 13.2 Hz, 3H), 3.79 (m, 3H), 4.15 (t, J ) 9.4 Hz, 3H),
4.35-4.51 (dd, J ) 4.1, 12.2 Hz, 6H), 4.49 (d, J ) 7.9 Hz, 3H),
4.52 (s, 2H), 4.78 (d, J ) 7.9 Hz, 3H), 5.25-5.37 (m, 6H), 5.61-
5.71 (m, 9H), 7.05-7.97 (m, 105H); 13C NMR (CDCl3, 100 MHz)
δ (ppm) 14.1, 30.0, 45.0, 60.8, 61.4, 62.8, 67.9, 68.0, 69.7, 70.3,
Supporting Information Available: Experimental procedure
for the synthesis of known compounds 8, 14, and 15. Copies of 1H
and 13C NMR spectra of compounds 2-7, 9a, 18-20, 22, 23, 25,
and 28. This material is available free of charge via the Internet at
http://pubs.acs.org.
JO071181R
J. Org. Chem, Vol. 72, No. 22, 2007 8279